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PhD project opportunities

On this page you can find out about PhD opportunities currently available in Biomedical Science. These projects cover developmental biology, cell biology, physiology and pharmacology, neuroscience, models of human disease, human stem cell biology and regenerative medicine.

Click on a project title below to find out more. Some of these projects come with funding to cover your tuition fees and living expenses, but they may not be available to students from outside the UK or the European Union.

Do you have your own idea for a project, or do you have your own funding in place?

Find a potential supervisor by visiting our research themes and centres webpages. Especially if you have your own funding in place we strongly encourage you to look at our science, and find a research area that excites you.

Contact a member of academic staff to find out about PhD opportunities in their area.

If you are applying for a project that does not come with funding, or you are not eligible for a funded project, there may be other ways for you to fund your PhD. We have a limited number of departmental scholarships available each year, and we also accept applications from students who are able to fund themselves.

It is a good idea to contact the supervisor of any PhD opportunity you want to apply for, before you submit your application. If the project you want to apply for does not come with funding, they may also be able to advise you on other sources of funding.

Once you have identified a project, a supervisor and a source of funding, you can complete the University's postgraduate online application form.


Fully funded projects

Developing novel biosensors for monitoring antibody production in CHO cells

Supervisor: Dr Andrew Peden

Funding status: This is a BBSRC-funded Industrial CASE PhD studentship and is full time for four years. The stipend will be £17,553.00 per annum. Applicant eligibility criteria for the studentship can be found at: www.bbsrc.ac.uk/documents/studentship-eligibility-pdf/ and you must identify that you fit these criteria prior to application.

Project Description

The biopharmaceutical market is valued at over 100 billion dollars per year with the majority of therapeutic antibodies being manufactured in Chinese Hamster Ovary (CHO) cells. However, the process of generating monoclonal antibody (mAb) producing cell lines suitable for commercial manufacture remains challenging, time consuming and costly.

The cellular pathways underpinning antibody production in CHO cells have been extensively studied over the past 10 years; however, it is still unclear which pathways are most important for this process (protein folding, protein transport and cell stress). The aim of this PhD is to facilitate a deeper understanding of CHO mAb production cell lines by investigating the intracellular phenotypes of CHO cell lines which are producing and secreting monoclonal antibodies. Using the knowledge gained we aim to define a phenotypic fingerprint which correlates with favourable manufacturing and product quality characteristics. Once these fingerprints have been defined, a series of novel fluorescent tools for monitoring them will be developed and applied to our cell line development platform to allow screening for optimal mAb producing cell lines in the earliest stages of cell line development.

This project will provide training in advanced mammalian cell culture, fluorescence microscopy, flow cytometry and access to state of the art, high-throughput, automated systems for cell culture (Berkeley Lights Beacon) and phenotypic analysis (Intellicyt iQue)   in place at the GSK laboratories.

The studentship is available starting in October 2017. This project is collaboration between Dr Andrew Peden at the University of Sheffield and Dr Robyn Emmins, GSK Biopharmaceutical Process Research, Stevenage. Candidates should have a strong academic degree in the biological sciences. The successful candidate will be highly motivated, work effectively in a team setting and be interested in cell biology, protein trafficking and technology development.

For informal enquiries about this project, please feel free to contact Andrew Peden

Web: Dr Andrew Peden

Email: a.peden@sheffield.ac.uk or Dr Robyn Emmins (robyn.a.emmins@gsk.com)

Molecular characterisation of the in vivo targets and mechanism of action of an acetylcholinesterase-derived peptide upregulated in Alzheimer’s disease - Funded by a four-year White Rose DTP studentship from the BBSRC.

Supervisor: Dr Mark Collins

Funding status: Funded by a four-year White Rose DTP studentship from the BBSRC to start in October 2017.

Project Description

Alzheimer’s disease (AD) is a progressive neurodegenerative disease that is characterised by the deposition and accumulation of amyloid beta plaques and phosphorylated tau filaments in the brain, the latter correlating with the onset of symptoms.

The vast majority of AD cases are sporadic and the basic mechanism driving the continuing process of neurodegeneration in selectively vulnerable cells, has not as yet been identified. However, our industrial partner, NeuroBio has recently determined that an AChE-derived 14-residue peptide (‘T14’) with a conspicuous sequence homology to amyloid beta, is upregulated in Alzheimer’s disease. T14 is present in all vulnerable cell populations irrespective of their neurotransmitter type and it drives the production of both amyloid and hyperphosphorylated tau.

These findings potentially place T14 upstream of amyloid and tau, however, a mechanistic understanding of the role of T14 in health and disease is lacking.

The aim of this project is to understand how T14 is involved in the pathogenesis of AD. We seek enthusiastic students who wish to gain expertise across disciplines including molecular neuroscience, protein biochemistry and mass spectrometry. The student will spend 6 months working in Neuro-Bio Ltd at their site in Abingdon, Oxfordshire.

Contact information:

For informal enquiries about this project, please feel free to contact Mark Collins.

Web: Dr Mark Collins

Email: mark.collins@sheffield.ac.uk

Competition and Self Funded Projects

Application deadline: Friday 30 June 2017

Elucidating the mechanisms of regulation of Nav1.7 channel in the plasma membrane of sensory neurons (Mohammed Nassar)

PhD Studentship starting October 2017

Title: Elucidating the mechanisms of regulation of Nav1.7 channel in the plasma membrane of sensory neurons

Supervisor: Dr Mohammed Nassar

Funding: Competition funded project UK students only

Project Description

"Sensory neurons detect and transmit painful stimuli to the CNS. Inflammation and nerve injury sensitise sensory neurons which results in a decrease of pain thresholds. This can be due, at least in part, to an enhanced trafficking of voltage gated sodium channels to the plasma membrane which would result in increased excitability of sensory neurons. The VGSC subunit Nav1.7 has been shown to be crucial for pain signalling in mouse and human. Three genetic pain disorders have been mapped to Nav1.7 in humans; these are primary erthromyalgia, familial rectal pain and complete insensitivity to pain. However, little is known about how the Nav1.7 surface pool is regulated to set pain thresholds and respond to changes in the environment (e.g. inflammation). Nav1.7 surface pool is determined by mechanisms controlling its transport to nerve terminals, insertion into and endocytosis from the membrane. Investigation of these processes may lead to new druggable targets for pain relief. The aim of this project is to identify the contribution of the intracellular parts of Nav1.7 channel in regulation of its membrane expression.

Methods work plan:
We have fused the reporter protein GFP to the Nav1.7 channel. This allows us to use live imaging to track Nav1.7 localisation in sensory neurons. Nav1.7 on plasma membrane will be quantified in standard and inflammation-like conditions. Live imaging will be performed using the DeltaVision OMX super resolution system. Published reports and our preliminary research indicated that the N and C-termini of Nav1.7 may play a role in regulating its surface pool.

DNA engineering will be used to produce N and C-termini mutants of the GFP-tagged channel to assess their contribution. Once the role of N and C-termini has been assessed, proteins that interact with them will be identified by a mass spectrometry. The role of the identified proteins will be validated by knockdown approaches. Lentiviruses will be used to introduce DNA coding for recombinant proteins and knockdown microRNA into sensory neurons. The project will involve using molecular biology methods to generate fusion protein constructs, transfection of DNA into cell lines and primary DRG neurons, immunocytochemistry and Western blotting. Functional effect of transfected fusions on Nav1.7 will be assessed by calcium imaging and patch clamping.

Impact:
The proposed work will provide a better understanding of Nav1.7 regulation in sensory neurons and may provide insights into pathways relevant to pathological changes in chronic pain conditions. Moreover, results may prove relevant to other membrane proteins in DRG whose surface pool could co-regulated with Nav1.7 by same pathways to decrease pain thresholds.

References

  1. Nociceptor-specific gene deletion reveals a major role for Nav1.7 (PN1) in acute and inflammatory pain. Nassar MA, Stirling LC, Forlani G, Baker MD, Matthews EA, Dickenson AH, Wood JN. Proc Natl Acad Sci U S A. 2004 Aug 24;101(34):12706-11.
  2. Veratridine produces distinct calcium response profiles in mouse Dorsal Root Ganglia neurons. Mohammed ZA, Doran C, Grundy D, Nassar MA. Sci Rep. 2017 Mar 24;7:
  3. Waxman SG, Merkies IS, Gerrits MM, Dib-Hajj SD, Lauria G, Cox JJ, Wood JN, Woods CG, Drenth JP, Faber CG. Sodium channel genes in pain-related disorders: phenotype-genotype associations and recommendations for clinical use. Lancet Neurol. 2014 Nov;13(11):1152-60.

For informal enquiries about this project, please contact:

Dr Mohammed Nassar

Email: m.nassar@sheffield.ac.uk

Exploring re-innervation and synaptogenesis between human stem cell-derived auditory neurons and inner hair cells: A therapeutic model for ‘hidden hearing loss’ (Marcelo Rivolta)

PhD Studentship starting October 2017

Title: Exploring re-innervation and synaptogenesis between human stem cell-derived auditory neurons and inner hair cells: A therapeutic model for ‘hidden hearing loss’

Supervisors: Professor Marcelo Rivolta and Dr Stuart Johnson

Funding: Competition funded project European/UK students only.

Project Description Loud noise exposure and aging lead to the permanent loss of afferent synaptic contacts to the primary sensory inner hair cells (IHCs) in the cochlea, whilst causing only a temporary or undetectable hearing threshold shift. This ‘synaptopathy’ reduces IHC afferent innervation by up to 50% leaving an individual unable to detect sounds in a noisy environment and experiencing difficulties with speech discrimination and intelligibility. Since the hearing thresholds are unaltered, it is difficult to detect the condition using conventional hearing tests, hence it is named ‘hidden hearing loss’. The loss of synaptic contacts is also thought to underlie primary neural degeneration in acquired hearing loss and aging. The causative mechanisms underlying cochlear synaptopathy are not well understood. While it is believed to be a result of the toxic effects of overstimulating these fibres, it is not known whether there are any changes in the properties of the IHCs that lead to axonal retraction or, conversely, whether there are any IHC changes that result from it.

Experimental Plan We have developed a way to make organotypic cultures of the excised mature mammalian cochlea and record from the sensory IHCs for 11 days in-culture. Interestingly, we found that after one or two days in culture the IHCs lose the physiological characteristics of mature cells and revert to a more immature phenotype without showing signs of degeneration or death. The PhD student will learn to make these cultures and use them to define the progression of this cell ‘de-differentiation’ using electrophysiological recording, staining and immunolabelling to see whether it is triggered by neuronal retraction or vice versa. We have previously shown that human embryonic stem cell-derived spiral ganglion neurons (SGNs) can be transplanted into an animal model and establish synaptic connections and recovery of function.

To address the specific role of the afferent fibres the student will learn to co-culture the adult gerbil organ of Corti with hESC-derived SGNs to see if the IHC properties have changed in order to attract new fibres and/or whether new synaptic innervation of the hair cells can restore their mature phenotype. This study will allow us to understand the chain of events that lead to the ‘de-differentiation’ of IHCs in culture and whether it can be reversed by re-innervation from stem cell-derived SGNs. The advantages of using the mature organotypic culture as a model system to study IHC and afferent fibre synaptopathy, and cochlear plasticity in general, are that the conditions can be easily manipulated whilst monitoring the characteristics of the IHCs and afferent fibres throughout.

Impact An understanding of cochlear plasticity in response to aging or trauma, and how to prevent or control it, will be essential for alleviating the effects of cochlear synaptopathy. This knowledge will also be important for slowing the progression of age-related hearing loss as well as the treatment of other poorly understood perceptual phenomena like tinnitus, the sensation of phantom tones, and hyperacusis, a reduced tolerance to moderate level sounds, all of which affect billions of people worldwide.

Keywords: Biophysics, Cell Biology / Development, Molecular Biology, Neuroscience / Neurology, Biophysics

References

  • Kujawa SG, Liberman MC (2015) Synaptopathy in the noise-exposed and aging cochlea: Primary neural degeneration in acquired sensorineural hearing loss. Hear Res 330: 191-199.
  • Johnson SL (2015) Membrane properties specialize mammalian inner hair cells for frequency or intensity encoding. eLife 4 pii: e08177
  • Chen W, Jongkamonwiwat N, Abbas L, Jacob Eshtan S, Johnson SL, Kuhn S, Milo M, Thurlow JK, Andrews PW, Marcotti W, Moore HD, Rivolta MN (2012) Restoration of auditory evoked responses by human ES-cell-derived otic progenitors. Nature 490: 278-282.

For informal enquiries about this project, please contact:

Can gut micro flora breakdown products be used as therapeutics in colorectal tumours?

Project Details

Supervisors - Dr Stephen Brown, Dr Bernard Corfe and Dr Paul Gokhale

The latest figures reveal there were 95,270 new cases of colon cancer in the United States (2016) and 41,265 in the UK (2014). Depending upon the country, it is either number 2 or 3 in the league table of cancer-related deaths and survival rates are low with 93% of stage 1 patients still alive after 5 years; stage 2, around 70-80%; stage 3, around 40-60%. Metastasis to the liver is the most common form of secondary tumours (~70%) and has the worst prognosis, with mean survival rates of approximately 9 months. The median time for liver metastases is about 18 months.

The Human intestinal epithelium is replaced every 4-6 days. The intestine possesses the stem cells, which differentiate into the cell types required for normal function. Colorectal cancer originates through mutations in the stem cells and is a multistage process, with the acquisition of mutations leading to the development of the disease. The process of the epithelial cell renewal is dependent upon the limited pool of multipotent intestinal stem cells.

The stem cells, which are maintained by continuous and rapid division in the crypts and subsequently differentiate into their progeny, is a highly controlled and coordinated process. Many, highly documented, signalling pathways are involved in their regulation. In combination with host factors, food and the Human Microbiome are also thought to play a role in homeostasis. The microbiome of the intestine can be varied and the metabolic products, potentially interesting for normal intestine health, could be of benefit as neutriceuticals in colorectal cancer.

The project is a collaboration between Dr. Bernard Corfe (Department of Oncology & Metabolism), who’s group investigate the relationship between diet, especially short-chain fatty acids, Dr. Paul Gokhale (Centre for Stem Cell Biology), who has interests in stem cell regulation and disease and Dr. Stephen Brown (primary supervisor), who’s research concerns organoid models and their use in drug discovery.

The project will investigate the role of microbial metabolites and how they change normal and diseased colorectal organoids. The PhD will involve organoid manipulation, high throughput technology, gene editing, gene expression studies, RNAi, computational data analysis and mining. The long-term goal of this project is to identify microbial breakdown products that can improve gut health or be used as colorectal cancer therapies.

Supervisors:

SRSF website: http://www.rnai.group.shef.ac.uk

If you are interested in any of the opportunities listed below, please contact the project supervisors directly to enquire about availability.

Investigating the role of the adhesion protein Tensin 3 in breast cancer invasion - Dr Elena Rainero

Supervisor: Dr Elena Rainero

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

The extracellular matrix (ECM) is a complex network of secreted proteins that, besides providing the scaffolding onto which tissue and organs are organised, is involved in the regulation of a variety of cell function, including survival, growth, migration and differentiation. Moreover, the interaction with the ECM has an important role in controlling tumour formation and metastasis in several cancer types, including breast cancer. Cells interact with the ECM through plasma membrane receptors, including the integrin family, which have a key role in controlling cancer cell invasion and migration. Integrins are not only found at the cell surface, but can be internalised and either be targeted for lysosomal degradation or transported back to the plasma membrane. Interestingly, the way in which integrins are trafficked intracellularly dictates how they control cell migration. My lab focuses on cell-ECM interaction from a novel and exciting angle, analysing the role of the internalisation of ECM components in promoting cancer invasiveness.

My data indicate that invasive breast cancer cells strongly up-regulate ECM uptake, compared to normal mammary epithelial cells, and the integrin-binding protein tensin-3 is required for this. Interestingly, high expression of tensin-3 correlates with poor prognosis in a cohort of breast cancer patients, supporting its role in cancer progression.

The aim of this project is the characterisation of the molecular mechanisms through which tensin-3 controls ECM internalisation and how this impinges on cancer cell invasion, analysing in particular: (1) whether tensin-3 controls ECM endocytosis through the regulation of integrin function; (2) how tensin-3 controls ECM organisation and turnover in 3D environment; (3) what is the role of tensin-3 in cancer cell migration and invasion, in both 2D and 3D settings. To mimic the physiological environment, fibroblast-generated cell-derived matrices and 3D culture systems will be used, coupled with confocal and time-lapse video microscopy.

The outcome of this project will shed new light on the role of ECM trafficking in cancer and will determine whether tensin-3 may represent a target for the development of novel anti-cancer therapeutic strategies.

Keywords: Cancer / Oncology, Cell Biology / Development

For informal enquiries about this project, please contact:

Role of primary cilia in skeletal muscle stem cells and muscle regeneration - Dr Anne-Gaelle Borycki

Supervisor: Dr Anne-Gaelle Borycki

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Primary cilia are ancient organelles present at the surface of many cell types in vertebrates. In recent years, they have been shown to play essential roles in relaying sensory and signalling information from the environment to the cell. The importance of primary cilia is underscored by the growing family of diseases associated with defects in cilia function, known as ciliopathies.

Satellite cells are skeletal muscle-specific stem cells responsible for the post-natal growth and repair following injury of skeletal muscles. Satellite cells are normally quiescent, but proliferate and differentiate to repair muscles when they become activated. We have previously shown that quiescent satellite cells exhibit primary cilia, which are rapidly disassembled upon satellite cell activation. Interestingly, we uncovered that in later phases of muscle regeneration, primary cilia re-assemble exclusively at the surface of self-renewing satellite cells and are essential for the maintenance of a stem cell pool in muscles.

This project aims at investigating further the role of primary cilia in satellite cell self-renewal and in muscle regeneration. The project will use a well-established ex-vivo culture system of skeletal muscle fibres as well as in vivo genetic approaches in vertebrates, imaging and molecular biology approaches to investigate whether primary cilia are essential cues for asymmetric cell division and stem cell self-renewal, and to establish the molecular mechanisms controlled by primary cilia in muscle regeneration.

References

  • Jaafar Marican NH, Cruz-Migoni SB, Borycki AG. (2016). Asymmetric Distribution of Primary Cilia Allocates Satellite Cells for Self-Renewal. Stem Cell Reports. 6(6):798-805.

Keywords: Cell Biology / Development, Genetics, Medical/Clinical Science, Molecular Biology

For informal enquiries about this project, please contact:

The role of extra-cellular matrix remodelling in muscle-specific stem cell activity - Dr Anne-Gaelle Borycki

Supervisor: Dr Anne-Gaelle Borycki

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Satellite cells are adult skeletal muscle-specific stem cells responsible for post-natal muscle growth and for muscle repair upon injury or disease such as dystrophies. Satellite cells are normally quiescent. However, upon activation, satellite cells exit quiescence, begin proliferating and initiate a myogenic program that culminates with the fusion to the injured muscle fibre and its repair. An important aspect of the repair program is the ability of satellite cells to self-renew, a process that ensures the maintenance of a stem cell pool throughout life allowing the repair of future injuries. Defects in satellite cell self-renewal underlie conditions such as muscular dystrophies and sarcopenia.

It is believed that the control of satellite cell quiescence, proliferation, differentiation and self-renewal is mediated through extrinsic signals produced by the satellite cell niche. We have previously shown that one such signal is delivered by the basal lamina associated with satellite cells, and is required for satellite cell self-renewal. This project aims at investigating further the mechanism of basal lamina control of satellite cell self-renewal. In particular, the project will address the relationship between the satellite cell basal lamina, cell polarity and asymmetric cell division, a process known to underlie self-renewal. The project will use a well-established ex-vivo culture system of skeletal muscle fibres, imaging and molecular biology approaches, combined with mouse genetics to gain a molecular and cellular understanding of satellite cell self-renewal.

Keywords: Cell Biology / Development, Genetics, Medical/Clinical Science, Molecular Biology

For informal enquiries about this project, please contact:

Cellular mechanisms underlying human infection by the intestinal bacterial pathogen enterohaemorrhagic Escherichia coli (EHEC) 0157 - Dr Daniel Humphreys

Supervisor: Dr Daniel Humphreys

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

The research group is funded by an MRC New Investigator Research Grant and a Department of Biomedical Science Start-Up Fund.

Understanding the molecular basis of host-pathogen interactions has never been more important given the health threat posed to humans and farmed food chain animals by bacterial pathogens that continue to develop multidrug resistance. Enterohaemorrhagic Escherichia coli (EHEC) 0157:H7 is an important enteric food-borne pathogen causing life-threatening haemorrhage colitis and haemolytic ureic syndrome in humans. Currently no treatment is available for EHEC infections. Advancing our knowledge of the EHEC virulence mechanisms has the potential to combat disease by speeding the development of anti-infectives and broadening the scope for therapeutic intervention.

To colonise the human intestine EHEC assembles a syringe-like Type 3 Secretion System that injects a cocktail of virulence effector proteins into host cells, which facilitate bacterial colonisation, survival and immune evasion in infected hosts (e.g. see Humphreys et al 2016). The effectors hijack cellular signalling pathways, including those involved in controlling the actin cytoskeleton, vesicle trafficking and cell survival. To combat this disease we need to know the host targets of the EHEC virulence effectors and the mechanisms by which they hijack cellular processes during infection.

We are looking for an enthusiastic and ambitious PhD candidate to investigate interactions between EHEC and mammalian host cells at the Department of Biomedical Science. The student will receive interdisciplinary training in a broad range of techniques, gaining valuable laboratory and research scientist experience, particularly in pathogen and cell biology, and will be an integral part of our research group and a collegiate Department.

The PhD project will combine pathogen genetics and molecular biology to engineer mutant strains of EHEC and recombinant virulence effectors. The recombinant strains will be used to identify pivotal virulence effectors and host cell targets (e.g. of the actin cytoskeleton, vesicle trafficking and/or cell survival) by performing high-throughput infection screens on cultured mammalian cells at the Sheffield RNAi Screening Facility. The screen will be used as a springboard to elucidate novel disease mechanisms by taking advantage of established protein-protein biochemical techniques, mass spectrometry, and cutting-edge fluorescence imaging microscopes at the Wolfson Imaging Light Microscopy Facility (e.g. super-structural resolution, confocal microscopy).

References

  1. Humphreys D. Singh V, Koronakis V. (2016). Inhibition of WAVE Regulatory Complex activation by a bacterial virulence effector counteracts pathogen phagocytosis. Cell Reports, in press.
  2. Humphreys D, Davidson AC, Hume PJ, Makin LE, Koronakis V. (2013) Arf6 coordinates actin assembly through the WAVE complex, a mechanism usurped by Salmonella to invade host cells. Proceedings of the National Academy of Sciences of the United States of America. 110(42):16880-16885. PMID: 24085844
  3. Humphreys D, Davidson AC, Hume PJ, Koronakis V. (2012) Salmonella SopE and host GEF ARNO cooperate to recruit and activate WAVE to trigger bacterial invasion. Cell Host & Microbe 11, 129-39. PMID: 22341462
  4. Smith, K., Humphreys, D., Hume, P.J., and Koronakis, V. (2010) Enteropathogenic Escherichia coli recruits the cellular inositol phosphatase SHIP2 to regulate actin-pedestal formation. Cell Host & Microbe 7, 13-24. PMID: 20114025

Keywords: Biochemistry, Cell Biology / Development, Medical/Clinical Science, Microbiology, Molecular Biology, Pathology

For informal enquiries about this project, please contact:

Mechanisms of cell signalling and coordination of cell polarity in animal development - Professor David Strutt

Supervisor: Professor David Strutt

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Tissue morphogenesis, repair and regeneration requires cells to communicate and coordinate their behaviours. Major pathways involved are the Wnt/Frizzled and Fat/Dachsous planar polarity pathways that mediate polarised cell signalling in epithelia sheets. Loss of their activity leads to a variety of developmental abnormalities in animal models such as failure of neural tube closure, cleft palates and heart defects, as well as deficits in wound healing and failure to repair kidney damage resulting in polycystic tubules, and is also implicated in cancer metastasis.

The project will focus on understanding molecular mechanisms of Wnt/Frizzled and Fat/Dachsous pathway activity, and in particular how individual proteins adopt polarised distributions with cells, and the signalling mechanisms that propagate this polarity from cell to cell. A molecular genetic approach will be used, taking advantage of the well-established model Drosophila, which provides sophisticated molecular and genetic and cell biological tools. A major focus in the lab is understanding protein dynamics during cell signalling, using techniques such as live imaging (both conventional and super-resolution) and FRAP, and combining this with genetic screens to identify new pathway components and cell biology and biochemical studies of protein behaviours. We combine these experimental studies with computation modelling approaches to aid in experimental design and hypothesis testing.

References

  • Hale, R., Brittle, A.L., Fisher, K.H., Monk, N.A. and Strutt, D. (2015) Cellular interpretation of the long-range gradient of Four-jointed activity in the Drosophila wing. eLife 4, e05789. [PMC4338440]
  • Strutt, H.*, Searle, E.*, Thomas-MacArthur, V., Brookfield, R. and Strutt, D. (2013) A Cul-3-BTB ubiquitylation pathway regulates junctional levels and asymmetry of core planar polarity proteins. Development 140: 1693-1702. [PMC3621487]
  • Brittle, A., Thomas, C. and Strutt, D. (2012) Planar polarity specification through the asymmetric subcellular localisation of the atypical cadherins Fat and Dachsous. Current Biology 22: 907-914. [PMC3362735]
  • Strutt, H.*, Warrington, S.J.* and Strutt, D. (2011) Dynamics of core planar polarity protein turnover and stable assembly into discrete membrane subdomains. Developmental Cell 20: 511-525. [PMC3094756]

Keywords: Biochemistry, Cancer / Oncology, Cell Biology / Development, Genetics

For informal enquiries about this project, please contact:

Cellular mechanisms underlying homeostatic balancing of neuronal excitation and inhibition in vivo - Dr Andrew Lin

Supervisor: Dr Andrew Lin

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Neuronal excitation and inhibition are very carefully balanced in the brain, and perturbed excitation/inhibition (E/I) balance has been linked to diseases such as epilepsy, autism and schizophrenia. Maintaining E/I balance within normal bounds depends in part on homeostatic plasticity, in which neurons compensate for deviations in activity levels by adjusting their responsiveness to excitation and inhibition. Although we are starting to understand the molecular mechanisms underlying homeostatic plasticity in reduced preparations, we still know very little about such mechanisms in the intact brain.

We have recently developed a new model system for addressing this question. In the fruit fly Drosophila, Kenyon cells (KCs), the neurons underlying olfactory associative memory, receive excitation from projection neurons as well as feedback inhibition from a single identified neuron. The balance between these two forces maintains sparse odour coding in Kenyon cells, which enhances the odour-specificity of associative memory by reducing overlap between odour representations. Preliminary evidence indicates that Kenyon cells adapt to prolonged disruption of E/I balance, providing a unique opportunity to use the powerful genetic tools of Drosophila to uncover the molecular mechanisms underlying homeostatic plasticity in the intact brain, in a defined circuit that mediates a sophisticated behaviour.

This project will test candidate cellular mechanisms underlying homeostatic compensation. For example, to compensate for insufficient inhibition onto Kenyon cells, excitatory synapses onto Kenyon cells might become weaker or smaller, or Kenyon cells might decrease their input resistance to become intrinsically less excitable. In testing whether these or other mechanisms underlie homeostatic plasticity in vivo, the student will develop skills in a wide range of techniques from fly genetics and confocal microscopy to patch-clamp electrophysiology, two-photon imaging of neural activity, and computational modelling.

References

  • About the model system: Lin, A.C., Bygrave, A.M., de Calignon, A., Lee, T., Miesenböck, G. (2014). Sparse, decorrelated odor coding in the mushroom body enhances learned odor discrimination. Nature Neuroscience, 17, 559-68.
  • Review about homeostatic plasticity: Davis, G. W. (2013). Homeostatic signaling and the stabilization of neural function. Neuron 80, 718–728.

Keywords: Bioinformatics, Cell Biology / Development, Genetics, Molecular Biology, Neuroscience/Neurology, Bioinformatics

For informal enquiries about this project, please contact:

Epithelial morphogenesis: coordinating planar polarity and tissue mechanics - Professor David Strutt

Supervisors:

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

As an organism develops, tissues are shaped and patterned in a coordinated way. The long-standing dogma of developmental biology is that secreted proteins diffuse to form expression gradients throughout tissues thereby providing spatial cues to direct growth, fate and pattern, drawing responses from cells even at some distance from the source. More recently, studies have revealed critical roles for mechanical forces in regulating morphogenesis, however, the interplay between these two systems is poorly understood. With the advent of fast 4D live imaging, combined with genetically encoded fluorescent sensors and sophisticated computational modelling tools, is it now possible to make major advances in understanding epithelial tissue dynamics at a quantitative systems level.

The model organism Drosophila provides an ideal system for dissecting mechanisms of morphogenesis in cell sheets, as it is highly amenable to genetic manipulation and has easily accessible simple tissues suitable for live imaging. The project will integrate cutting-edge genetic tools, advanced 4D fast live imaging and computational modelling in an iterative manner to: (i) explore how mechanical forces influence patterning and polarity; (ii) understand how cell division modulates tissue mechanics and coordinated cell polarity; and (iii) develop mathematical approaches to incorporate pattern and proliferation within existing modelling frameworks for epithelial morphogenesis.

We are looking for an enthusiastic and ambitious student to carry out this interdisciplinary project using both experimental and computational approaches. This project will be suitable for a student with a strong quantitative background (e.g. mathematics, physics, engineering or computer science) who is keen to apply their skills to a biological problem with potentially significant translational importance, or a student with a biological background but a strong interest and some skills in computational approaches. Moreover, the student will be provided with an interdisciplinary training, gaining valuable laboratory experience, particularly in high-level imaging and image analysis, as well as in mathematics and computational modelling.

References

  • Warrington SJ, Strutt H and Strutt D. (2013) The Frizzled-dependent planar polarity pathway locally promotes E-cadherin turnover via recruitment of RhoGEF2. Development 140(5):1045-1054.
  • Strutt H*, Warrington, SJ* and Strutt D. (2011) Dynamics of core planar polarity protein turnover and stable assembly into discrete membrane subdomains. Developmental Cell 20: 511-525.
  • Kursawe J, Brodskiy PA, Zartman JJ, Baker RE, Fletcher AG. Capabilities and Limitations of Tissue Size Control Through Passive Mechanical Forces. PLoS Comput Biol. 2015 Dec 29;11(12):e1004679.
  • Tetley RJ, Blanchard GB, Fletcher AG, Adams RJ, Sanson B. Unipolar distributions of junctional Myosin II identify cell stripe boundaries that drive cell intercalation throughout Drosophila axis extension. eLife. 2016 May 16;5:e12094.

Keywords: Cell Biology / Development, Genetics, Applied Mathematics

For informal enquiries about this project, please contact:

Host-pathogen interactions of the human fungal pathogen Cryptococcus neoformans - Dr Jason King

Supervisors:

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

This project will investigate how the fungal pathogen Cryptococcus neoformans interacts with host cells (https://goo.gl/bJaQco). Cryptococcal infection is both a significant opportunistic infection that causes hundreds of thousands of deaths worldwide each year and an excellent opportunity to understand how pathogens avoid killing by the immune system.

Like many opportunistic pathogens Cryptococcus has not evolved specifically to avoid human immune cells. Rather, the normal environmental niche is soil, where its primary concern is capture by predatory amoeba. This project will therefore take a novel approach and use the amoeba Dictyostelium discoideum to investigate what happens to Cryptococcus within a physiologically revelant, model phagocytic cell. This provides an outstanding model system that allows us to genetically manipulate both the host and pathogen, and follow the transit of Cryptococcus through its host in great detail using time lapse fluorescence microscopy. This will allow the successful student to dissect how host and pathogen interact in greater detail than previously possible.

The primary aim of this study is to understand what happens to the human pathogen Cryptococcus neoformans after ingestion by phagocytic cells. This is the first line of defence in the immune system, and the ability of Cryptococcus to manipulate host phagocytes and evade digestion is the major cause of its pathogenicity (goo.gl/IkIf2V). Amazingly Cryptococcus is able to escape macrophages by a process called vomocytosis (goo.gl/BVKzL5). We have shown that Cryptococcus is also able to avoid being killed by Dictyostelium, as well as escape via vomocytosis.

This project with therefore use a combination of genetics, live cell imaging, biochemistry and flow cytometry to identify and characterize how Cryptococcus manipulates its host. This will provide a new level of understanding of cryptococcosis and identify new avenues for therapeutic intervention.

Keywords: Biochemistry, Cell Biology / Development, Genetics, Immunology, Medical/Clinical Science, Microbiology, Molecular Biology

For informal enquiries about this project, please contact:

Please see the King and Johnson lab websites for more details, and recent publications:

The role of the intracellular parts of the Nav1.7 channel in its membrane expression - Dr Mohammed Nassar

Supervisor: Dr Mohammed Nassar

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Sensory neurons detect and transmit painful stimuli to the CNS. Inflammation and nerve injury sensitise sensory neurons in vivo which results in a decrease of pain thresholds. This can be due, at least in part, to an enhanced trafficking of voltage gated sodium channels to the plasma membrane which would result in increased excitability of sensory neurons. The VGSC subunit Nav1.7 has been shown to be crucial for pain signaling in mouse and human.

Three genetic pain disorders have been mapped to Nav1.7 in humans; these are primary erthromyalgia, familial rectal pain and complete insensitivity to pain. However and despite its importance in pain signaling, little is known about Nav1.7 transport to nerve terminals, insertion into the membrane and endocytosis from the membrane. Investigation of these processes may lead to new druggable targets for pain relief. Drugs will be directed to inhibit insertion in membrane and increase endocytosis form membrane. Therefore, the aim of this project is to identify the contribution of the intracellular parts of Nav1.7 channel to its membrane expression.

The project will involve using molecular biology methods to generate fusion protein constructs, transfection of DNA into cell lines and primary DRG neurons, immunocytochemistry and Western blotting. Functional effect of transfected fusions on Nav1.7 will be assessed by calcium imaging and patch clamping.

For further details and informal discussion please feel free to contact Dr Nassar.

Keywords: Cell Biology / Development, Molecular Biology, Neuroscience/Neurology

For informal enquiries about this project, please contact:

Elucidating the cellular pathways and machinery involved in epithelial polarisation and cancer using advanced bioimaging - Dr Andrew Peden

Supervisors:

This project is eligible for funding. Find out more on our funding webpage.

Epithelial polarisation plays an important role in tissue morphogenesis and perturbations in this process can lead to developmental defects and cause cancer. Approximately, 90% of all cancers are derived from epithelial tissue (carcinomas). My group has recently identified a novel vertebrate- and epithelial-specific SNARE, STX19, that is required for the delivery of biosynthetic and endocytic material to the cell surface. Using proteomics and yeast-two-hybrid screening we have identified several novel, STX19 interacting partners that are important players in establishing and maintaining epithelial polarity. It is our hypothesis that STX19 plays a role in the delivery of cell polarity proteins to the cell surface and is required for epithelial polarisation.

To test this hypothesis shRNA will be used to knock down STX19 and its interacting partners in 2 and 3D cell cultures models and epithelial polarisation visualized using structured illumination microscopy. To assess the physiological role of STX19, CRISPR/Cas9 will be also used to generate a STX19 knockout zebrafish and organ development monitored using light sheet microscopy. Using both SIM and light sheet microscopy will allow the processes that drive tissue development to be visualised at an unprecedented spatiotemporal resolution.

This project represents an exciting training opportunity where the student will become an expert in advanced bioimaging, cell biology and animal physiology. This project will not only provide insight into the fundamental cellular processes governing epithelial polarisation and tissue development but also shed light on how changes in these pathways can cause cancer.  

Keywords: Biochemistry, Cancer / Oncology, Cell Biology / Development, Molecular Biology

For informal enquiries about this project, please contact:

Obtaining a molecular understanding of antibody secretion - Dr Andrew Peden

Supervisor: Dr Andrew Peden

This is a self funded project. Find out more on our funding webpage.

Plasma cells are the antibody producing cells of the immune system so are vital for fighting infection.  Plasma cells secrete hundreds of millions of antibody molecules per day and this is achieved through a dramatic upregulation of the biosynthetic pathway.

Dysregulation of this process has been implicated in a wide range of diseases from benign autoimmune disorders to metastatic cancer. The aim of this project is to identify novel factors required for regulating antibody secretion and plasma cell function. These factors will be identified using a combination of comparative transcriptomics, proteomics and functional genomics. Genes identified using these methods will increase our fundamental understanding of the secretory pathway and may also be useful for modulating plasma cell function.

The PhD candidate will be trained in techniques such as primary cell culture, proteomics and high-throughput screening.

Keywords: Biochemistry, Bioinformatics, Biotechnology, Cancer / Oncology, Cell Biology / Development, Immunology, Molecular Biology, Bioinformatics

For informal enquiries about this project, please contact:

Cleavage of the protrusion regulator, Vav2, directs cell migration during wound healing - Dr Mark Bass

Supervisor: Dr Mark Bass

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Healing delays affect 200,000 UK patients a year, often result in limb amputation, and are caused by defects in the migration of skin fibroblasts.  Upon injury to healthy skin, resident skin fibroblasts are activated and cluster at the wound shoulder where they contract the defect.  Failures in fibroblast migration cause healing delays to the extent that fibroblast senescence is one of the two hallmarks of chronic wounds.  Therefore understanding how migration of fibroblasts towards wound signals is regulated is critical.  The cytoskeletal regulator, Rac1, both drives and guides migration by stimulating localised membrane protrusion. 

A key question is how fibroblasts recognise wound signals and polarise protrusion so that migration to the wound site is efficient.  Rac1 itself is activated by exchange factors such as Vav2, in response to extracellular changes in the wound environment.  We have preliminary evidence that Vav2 mediates the migratory response during healing and that Vav2 is cleaved, immediately after activation, thus ensuring that protrusion signals are transient, making cells responsive to changing migration cues.  This project will identify the Vav2 cleavage site and determine how that cleavage event affects decision making as fibroblasts migrate.

This project will use mass spectrometry to map the site of Vav2 cleavage in response to wound signals, so that cleavage-resistant mutants and pre-cleaved fragments can be generated.  The mutants will be used to investigate the effect of cleavage on Rac1 regulation in cell-based assays and FRET experiments using Rac1 activity probes.  Following characterisation of the effect of cleavage on Rac1, the mechanism will be investigated, thus determining how Vav2 regulation is linked to the extracellular events that occur upon wounding.  Of critical importance will be the investigation of how Vav2 cleavage affects migration.  3D matrices that resemble the skin will be generated and used to investigate the effect of Vav2 cleavage on migration persistence, while micropatterned surfaces will be used to test the effect on decision-making when faced with alternative paths during migration.  Together these experiments will resolve the mechanism by which protrusion is regulated and migration directed during healing.

Finally, the project will link to our ongoing clinical investigations.  We are in the process of developing ultrasonic therapies that restore wound closure in healing-defective animals and patients.  The major limitations to clinical translation of the work are the gaps in our understanding of how Rac1 activation is regulated during healthy healing.  The progress in this PhD project will aid in predicting the healing prognosis of patients, including identification of groups that need and would respond well to intervention.  The effect of the Vav2 cleavage mutants on Rac1 activation in response to ultrasonic stimuli will be tested, thus ensuring that the project connects molecular mechanism to therapeutic outcome and that the student gains understanding of therapeutic development.

References

Roper, Williamson, Bally, Cowell, Brooks, Stephens, Harrison, Bass. (2015) Ultrasonic stimulation of mouse skin reverses the healing delays in diabetes and aging by activation of Rac1. J Invest Dermatol. 135: 2842. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4902130/

Williamson, Hammond, Bergen, Roper, Feng, Rendall, Race, Bass. (2014) A coronin-1C/RCC2 complex guides mesenchymal migration by trafficking Rac1 and controlling GEF exposure. J Cell Sci. 127: 4292. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4179493/

Bass, Williamson, Nunan, Humphries, Byron, Morgan, Martin, Humphries. (2011) A syndecan-4 hair trigger initiates wound healing through caveolin- and RhoG-regulated integrin endocytosis. Dev Cell. 21: 681. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC3202633/

Keywords: Biochemistry, Cancer / Oncology, Cell Biology / Development, Molecular Biology

For informal enquiries about this project, please contact:

Localisation of Rac1 signals in chronic wound and cancer-associated fibroblasts - Dr Mark Bass

Supervisor: Dr Mark Bass

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Healing defects are one of the largest current health challenges.  Chronic leg ulcers affect 200,000 UK patients and cost the NHS £3.1 billion (3% of expenditure) annually, and the rise in linked risk factors such as age and diabetes mean that the challenge is increasing.  Although not fatal, chronic wounds have a huge effect on quality of life as they cause chronic pain and frequently result in amputation of the limb.  Therefore if we are to achieve lifelong health, it is essential that we understand the healing process and identify ways to reverse the defect when healing breaks down.  In healthy individuals, dermal fibroblasts migrate to the wound bed, where they contract to draw the edges of the wound together, but in unhealthy individuals, defects in fibroblast migration are one of the two hallmarks of chronic wounds.

Our laboratory investigates the regulation of migratory signals induced by changes in the extracellular matrix that occur upon wounding.  Directional fibroblast migration requires: a) activation of the protrusion signal, Rac1, at the front of the cell, but suppression at the rear, b) precise temporal regulation of protrusion signals to allow coordination with cell contraction to pull the cell body forward.  We recently reported a Rac1-sequestering molecule, RCC2, and newer data have revealed a link between RCC2 and the transmembrane matrix sensor, syndecan-4.  Genetic disruption of either RCC2 or syndecan-4 perturbs the directionality of fibroblast migration, and both have been demonstrated in vivo.  RCC2-depletion in zebrafish compromises developmental migration, while syndecan-4-knockout in mice retards wound healing.  Both have also been linked to cancer development, and whether this is due to a change in the cancer cells themselves, or cancer-associated fibroblasts remains to be seen.

This project will determine how signalling from syndecan-4 to RCC2 localises Rac1 signalling and thereby directs fibroblast migration.  The composition of the RCC2 complex will be determined by combining syndecan-4 and RCC2 mutants with knockdown of other complex components, identified by mass spectrometry.  The influence of those interactions on Rac1 signalling will be determined using cell-based signalling assays and FRET.  Crucially you will test the effect of the pathways on fibroblast migration through synthetic skin mimics and genuine skin sections.

This project will also link to our ongoing investigations into the use of ultrasound to promote Rac1 activation in cells where matrix-dependent signalling is defective.  We have demonstrated that ultrasound can overturn healing defects in diabetic and syndecan-4-knockout mice and are now testing whether it reverses senescence in both chronic wound and cancer-associated fibroblasts.  The putative role of the syndecan-4/RCC2 pathway in both these instances suggests a link to ultrasound efficacy, meaning that this project will result in clinical impact within a few years.

References

Roper, Williamson, Bally, Cowell, Brooks, Stephens, Harrison, Bass. (2015) Ultrasonic stimulation of mouse skin reverses the healing delays in diabetes and aging by activation of Rac1. J Invest Dermatol. 135: 2842. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4902130/

Williamson, Hammond, Bergen, Roper, Feng, Rendall, Race, Bass. (2014) A coronin-1C/RCC2 complex guides mesenchymal migration by trafficking Rac1 and controlling GEF exposure. J Cell Sci. 127: 4292. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4179493/

Bass, Roach, Morgan, Mostafavi-Pour, Schoen, Muramatsu, Mayer, Ballestrem, Spatz, Humphries. (2007) Syndecan-4-dependent Rac1 regulation determines directional migration in response to the extracellular matrix. J Cell Biol. 177:527. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC1885470/

Keywords: Biochemistry, Cancer / Oncology, Cell Biology / Development, Molecular Biology

For informal enquiries about this project, please contact:

Integrity of mitochondrial DNA in human pluripotent stem cells and implications for regenerative medicine - Dr Ivana Barbaric

Supervisor: Dr Ivana Barbaric

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Human Pluripotent Stem Cells (hPSCs) have the ability to produce cells of all tissues within the body. They can provide a valuable tool for modelling human disease, as well as a potential source of differentiated cells for use in regenerative medicine and drug discovery. In order to fulfil the therapeutic potential of hPSCs it is important to ensure their genome is free of any potentially deleterious mutations that could either impair the function of the hPSC-derived differentiated derivatives or give rise to malignant phenotypes upon transplantation in vivo. Recurrent genetic changes have been noted in the nuclear genome of hPSCs that appear to confer selective growth advantage to the variant cells. Less understood is the appearance and role of mutations in the mitochondrial genome of hPSCs.

This project will focus on investigating the extent of mitochondrial DNA instability, molecular mechanisms that underpin mutation generation and the functional consequences of the mutations on hPSC phenotype and behaviour. Techniques will include a range of molecular and cellular biology methods, including hPSC culture, genetic manipulation of hPSC using CRISPR/Cas9, flow cytometry and live cell imaging.

Keywords: Cell Biology, Development.

For informal enquiries about this project, please contact:

Imaging life beyond the diffraction limit of light - Dr Jarema Malicki

Supervisor: Dr Jarema Malicki

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Ever since its invention centuries ago, light microscopy has been hampered by the diffractive properties of light, which limit resolution to ca. 200 nm.  This resolution limit precludes the visualization of many subcellular structures using light microscopy and has been a major obstacle in the imaging of biological processes.  Recently, advances in fluorophore excitation methods and image processing algorithms have overcome this limitation, increasing image resolution by as much as 10 times to about 20 nm.  This new form of imaging is termed super-resolution microscopy.

Super-resolution microscopy opens unprecedented opportunities for imaging biological structures.  We take advantage of this approach to image subcellular structures that regulate intracellular traffic.  We focus on a barrier structure that regulates protein movement between the cell’s cytoplasm and the cilium, a tiny subcellular compartment on the surface of cell.  The cilium is just 250 nm across and so conventional light microscopy cannot be used to visualize its inner architecture.  The inner components of cilia are, however, essential for the function of many cells, tissues, and organs.

We have recently obtained excellent quality super-resolution images in a simple unicellular organism, Tetrahymena.  The purpose of this project is to extend these imaging studies to vertebrate tissues, focusing on the nervous system.  To this end, we will use transgenic lines that express specialized fluorescent proteins suitable for super-resolution imaging in sensory neurons.  This will make it possible to image vital structures of these neurons, such as cilia or synaptic termini, in unprecedented detail and thereby gain insight into the function of these cells.

Keywords: Cell Biology/ Development, Genetics, Neuroscience/Neurology

For informal enquiries about this project, please contact:

or see the Malicki lab website: http://www.malickilab.org/

Developing drugs for long-lasting pain relief - Professor Bazbek Davletov

Supervisor: Professor Bazbek Davletov

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

This project aims to develop new approach for long-lasting pain relief. Around 12% of adults suffer from severe chronic pain which include cancer, inflammatory and neuropathic pain. Available drugs to treat persistent pain are rarely curative and bring intolerable side effects in a long run. A key feature of our approach is the use of re-targeted botulinum proteases to selectively silence specific types of neurons for months-long periods of time after local injections.

This strategy has evolved from my studies of neuronal communication which universally depends on SNARE proteins. We and others demonstrated that specific cleavage of these proteins can lead to prolonged silencing of neurons with full recovery after several months. My laboratory recently engineered botulinum molecules which have a more selective action. Specifically, several of our molecules target central and sensory neurons but not neuromuscular junctions. This feature makes novel botulinum molecules attractive in treating various chronic neuronal disorders since neuronal silencing can be achieved without side-effects.

We plan to obtain evidence in cell cultures that these botulinum molecules can block release of pain-signalling molecules from sensory neurons and then to test them for their ability to cause long-lasting analgesia in rodent models. Proving the analgesic potential of specific botulinum drugs will be required to translate the benefits of basic research to clinical practice.

References

Mangione AS, Obara I, Maiarú M, Geranton SM, Tassorelli C, Ferrari E, Leese C, Davletov B, Hunt SP. ‘Non-paralytic botulinum molecules for the control of pain. Pain 157:1045-55.

Ferrari E, Gu C, Niranjan D, Restani L, Rasetti-Escargueil C, Obara I, Geranton SM, Arsenault J, Goetze TA, Harper CB, Nguyen TH, Maywood E, O'Brien J, Schiavo G, Wheeler DW, Meunier F, Hastings M, Edwardson JM, Sesardic D, Caleo M, Hunt SP & Davletov B ‘A synthetic self-assembling clostridial chimera for modulation of sensory functions’ Bioconjug Chem 24, 1750−1759.

Arsenault J, Ferrari E, Niranjan D, Cuijpers SAG, Gu C, Vallis Y, O'Brien J & Davletov B. ‘Stapling of the botulinum type A protease to growth factors and neuropeptides allows selective targeting of neuroendocrine cells’ J Neurochem 126, 223-33.

Keywords: Biochemistry, Biotechnology, Cell Biology / Development, Molecular Biology, Neuroscience/Neurology, Pharmacology, Veterinary Medicine, Macromolecular Chemistry, Pharmaceutical Chemistry

For informal enquiries about this project, please contact:

Establishment of an in vitro model for cystic fibrosis using organ on chip technology - Dr Kai Erdmann

Supervisor: Dr Kai Erdmann

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Cystic fibrosis is a devastating disease affecting the lung and the intestine. Cystic fibrosis is a genetic disease and caused by mutations of the chloride channel CFTR (cystic fibrosis transmembrane conductance regulator). Major phenotypes are increased mucus production in the lung causing extreme breathing difficulties as well as defective signaling in the intestine. One in three thousand newborns are affected.

The goal of this PhD project is the establishment of a state of the art in vitro model for cystic fibrosis to study the disease mechanism in more detail but also to establish a model suitable for high-throughput drug screening. The project will make use of modern organ on chip technology, which allows the functional reconstitution of organ functionality in miniaturized microfluidic devises. Precisely we will apply devices to mimic blood and airflow of the lung allowing a more physiological model system than just 2 or 3D cell cultures.

The aims of this project are
1.) to develop a cellular lung and/or gut model for cystic fibrosis that recapitulates the  phenotype observed in cystic fibrosis.
2.) to incorporate this cellular model into an organ on a chips suitable for mechanistic investigations as well as for high throughput screening
3.) to perform experiments demonstrating suitability of this organ on a chip model for high throughput screening to identify drugs and to analyse the CFTR underlying disease mechanism in more detail.

Techniques: CRISPR-technology, organoids, 3D cell cultures with different mechanical properties, micro-molding using soft-litography, Micropatterning, siRNA-technology, Cell imaging (live confocal spinning disk and super-resolution microscopy, Total-internal reflection microscopy)

References

  • Cutting, G.R. Cystic fibrosis genetics: from molecular understanding to clinical application Nature Reviews Genetics 16, 45-56 (2015)
  • Guggino, W.B. & Stanton, B.A. Mechanisms of disease: New insights into cystic fibrosis: molecular switches that regulate CFTR Nature Reviews Molecular Cell Biology 7, 275-283 (2006)
  • Huh, D. et al. Reconstituting organ-level lung functions on a chip. Science 328, 1662–1668 (2010).

Keywords: Biochemistry, Biotechnology, Cell Biology / Development, Molecular Biology, Neuroscience/Neurology, Pharmacology, Veterinary Medicine, Macromolecular Chemistry, Pharmaceutical Chemistry

For informal enquiries about this project, please contact:

The role of multi-PDZ domain proteins in the regulation of epithelial cell proliferation and cancer - Dr Kai Erdmann

Supervisor: Dr Kai Erdmann

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

PTPN13 is a highly modular multi-PDZ domain protein composed of an N-terminal KIND (Kinase non catalytic C-lobe) domain, followed by a FERM (Four-point-one/Ezrin/Radixin/Moesin) domain, five PDZ domains, and a tyrosine phosphatase domain assembling a large macromolecular protein complex. There is strong evidence that PTPN13 plays a role in tumor development. In a search for protein tyrosine phosphatases involved in formation of colon carcinoma, it was demonstrated that PTPN13 is mutated in around 10% of colon carcinomas. We have recently identified novel components of the PTPN13 complex. This complex plays an important role in the regulation of cell/cell adhesion and nuclear transcription. However, the precise molecular mechanism, how PTPN13 contributes to tumor development is largely unclear

The goal of this project is to investigate the role of PTPN13 and other multi-PDZ domain proteins in the context of contact inhibition of proliferation, metastasis and tumor invasion. We will apply in vitro cell proliferation, migration and tumor invasion assays as well as state of the art still- and live-imaging microscopy techniques (like spinning disc confocal microscopy and superresolution microscopy) to investigate underlying disease mechanisms.  Furthermore, using in vitro 3-dimensional cell culture systems and modern chip based cell micropatterning we will further analyze the role and molecular mechanism of PTPN13 in epithelial cell proliferation and cell polarization.

References

  •  Hagemann, N., Ackermann, N., Christmann, J., Brier, S, Yu, F., Erdmann, K.S. The serologically defined colon cancer antigen-3 interacts with the protein tyrosine phosphatase PTPN13 and is involved in the regulation of cytokinesis. Oncogene, 32, 4602-4613 (2013)
  • Stenzel, N., Fetzer, C.P., Heumann, R., Erdmann, K.S. PDZ-domain directed basolateral targeting of peripheral membrane proteins in epithelial cells. J. Cell Sci., 122, 3374-3384 (2009)
  • Freiss, G., Chalbos, D. PTPN13/PTPL1: an important regulator of tumor aggressiveness. Anticancer Agents Med Chem. 11, 77-88 (2011)

Keywords: Biochemistry, Cancer / Oncology, Cell Biology / Development, Medical/Clinical Science, Molecular Biology

For informal enquiries about this project, please contact:

Developing a novel methotrexate-derived small molecule JAK/STAT pathway inhibitor - Dr Martin Zeidler

Supervisors:

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Work in the Zeidler lab has previously identified the anti-folate drug methotrexate (MTX) as a potent JAK/STAT pathway inhibitor. Used at low doses for the treatment of inflammatory and autoimmune conditions such as rheumatoid arthritis, methotrexate is widely used, has a well understood toxicity profile and is a very low cost drug. Given its function as a JAK/STAT inhibitor, the potential exists to repurpose MTX for other diseases associated with inappropriate pathway activity. Such a change in clinical practice has recently been demonstrated and has significant potential to fulfill a significant unmet need.

However, while repurposing MTX has considerable potential, its long term use is also sometimes associated with potentially serious side-effects – most of which are the consequence of its inhibition of dihydrofolate reductase activity (DHFR). MTX side effects range from relatively minor gastro-intestinal discomfort and nausea to potentially serious myelosuppression and foetal loss/defects in early pregnancy. Given these undesirable characteristics, the development of novel MTX-derived compounds which retain their ability to suppress the JAK/STAT pathway, but which no longer inhibit DHFR, have the potential to define a new class of ‘next generation’ JAK/STAT pathway inhibitors.

In this project, cell based assays for JAK/STAT and DHFR activity already established in the Zeidler lab will be combined with novel medicinal chemistry approaches in the Partridge lab. The successful candidate will design and synthesize libraries of novel MTX derivatives designed to explore the chemical space around methotrexate. The candidate will then use these compounds, together with established positive and negative controls, to undertake cell based assays designed to establish the specificity, activity and toxicity of the molecules generated. Lead compounds will be refined using iterative ‘molecular evolution’ approaches and established medicinal chemistry techniques. Ultimately, compounds will be tested in pre-clinical Drosophila and mouse models of human JAK/STAT disease. In the long-term, we aim to engage with the pharmaceutical industry as a prelude to potential human clinical trials.

References:

  • Thomas S, Fisher KH, Snowden JA, Danson SJ, Brown S, Zeidler MP (2015) “Methotrexate is a JAK/STAT pathway inhibitor” PLOSone 10(7) e0130078
  • Palandri, Francesca; Labate, Claudia; Sabattini, Elena; et al. (2016) “Low-dose methotrexate as treatment of myeloproliferative neoplasms: Proof of principle of clinical activity” Am J HEMATOLOGY 91(8) E329-E330

Keywords: Biochemistry, Cancer / Oncology, Medical/Clinical Science, Neuroscience/Neurology, Organic Chemistry, Pharmaceutical Chemistry

For informal enquiries about this project, please contact:

SUMOylation regulation of autophagy induced by ER stress in motor neuron disease - Dr Chun Guo

Supervisor: Dr Chun Guo

Co-supervisor: Professor Matthew Holley

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Autophagy is essential for cellular homeostasis and survival. Dysfunctional autophagy has been detected in various neurodegenerative disorders, including Alzheimer's disease (AD) and Amyotrophic lateral sclerosis (ALS)(1), and it is regarded as a suitable target for therapeutic intervention. It leads to stress in the endoplasmic reticulum (ER), which triggers the unfolded protein response (UPR) (2). The UPR has three canonical arms: IRE1α (the inositol-requiring kinase 1 α), PERK and ATF6. Abnormal activation of the IRE1α-XBP1 pathway has been detected in AD and ALS and inhibition of IRE1α activity increases cell viability in ER stress-induced degeneration (3). One function of XBP1 is to suppress neuronal autophagy and its depletion promotes autophagy, enhancing clearance of protein aggregates and ameliorating disease progression in ALS (4). Emerging evidence suggests an important role for protein SUMOylation in UPR, which influences cell survival (5). SUMOylation of XBP1 negatively regulates its function(6), so it is tempting to speculate that it can promote autophagy upon ER stress.

This exciting project will test the hypothesis that protein SUMOylation can reduce disease pathology in inherited forms of ALS by restoring autophagy via the IRE1α-XBP1 pathway. The proposed work will be conducted using a combination of techniques including molecular biology (e.g. cloning, tagging, mutagenesis), protein chemistry (e.g. GST-/His-pulldowns, co-immunoprecipitations (co-IPs), protein purification for assays of SUMOylation and protein interaction, western blotting for the detection of LC3 conversion from LC3-I to LC3-II and degradation of p62), cell culture (clonal cell lines, motor neuronal cell lines and cells derived from ALS animal models or human patients), image analysis of changes in cell/tissue morphology/histology, protein aggregation and ‘autophagic flux’ (e.g. immunocytochemistry, fluorescence and confocal microscopy) and cell viability (e.g. cytochrome c release, caspase activation, MTT, LDH assays).

References:

  1. Nixon, R. A. The role of autophagy in neurodegenerative disease. Nature medicine 19, 983-997, doi:10.1038/nm.3232 (2013).
  2. Scheper, W. & Hoozemans, J. J. The unfolded protein response in neurodegenerative diseases: a neuropathological perspective. Acta neuropathologica 130, 315-331, doi:10.1007/s00401-015-1462-8 (2015).
  3. Ghosh, R. et al. Allosteric inhibition of the IRE1alpha RNase preserves cell viability and function during endoplasmic reticulum stress. Cell 158, 534-548, doi:10.1016/j.cell.2014.07.002 (2014).
  4. Hetz, C. et al. XBP-1 deficiency in the nervous system protects against amyotrophic lateral sclerosis by increasing autophagy. Genes & development 23, 2294-2306, doi:10.1101/gad.1830709 (2009).
  5. Guo, C. & Henley, J. M. Wrestling with stress: roles of protein SUMOylation and deSUMOylation in cell stress response. IUBMB life 66, 71-77, doi:10.1002/iub.1244 (2014).
  6. Chen, H. & Qi, L. SUMO modification regulates the transcriptional activity of XBP1. The Biochemical journal 429, 95-102, doi:10.1042/bj20100193 (2010).

Keywords: Biochemistry, Cell Biology / Development, Genetics, Medical/Clinical Science, Molecular Biology, Neuroscience/Neurology, Pathology, Pharmacology, Statistics

For informal enquiries about this project, please contact:

Regulation of interphase microtubules by actin cytoskeleton - Dr Natalia Bulgakova

Supervisor: Dr Natalia Bulgakova

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Intracellular arrangement of microtubule cytoskeleton in interphase cells is diverse across tissues and cell types, ranging from radial patterns to parallel arrays. Aligned microtubules are a hallmark of specialized cell types with non-centrosomal microtubules, such as neuronal and epithelial cells. The microtubule alignment creates a structural scaffold for vectorial transport of different cargos, and therefore is crucial for cell polarity, cell shape, cell migration, and cell-cell communication.

Recently, we found that alignment of microtubules in epithelial cells is largely influenced by two factors. On the one hand, it is achieved by the response of growing microtubules to the geometric constraints of the cell. On the other hand, not all properties of microtubule organisation can be explained by cell geometry. Another type of cytoskeleton, actin, influences arrangement of microtubules, and therefore, their function in epithelial cells. We found that depolymerisation of actin cytoskeleton significantly changes microtubule alignment in Drosophila epithelial cells.

This project will address two related questions:

  1. What is the molecular mechanism of microtubule regulation by actin?
  2. What is the contribution of this regulation to the development of an organism?

During the project progression, the student will receive training in a wide range of techniques including molecular biology, state-of-art microscopy (live imaging, super-resolution) and computational approaches. This project will be done in collaboration with applied mathematicians from the group of Dr. Lyubov Chumakova, University of Edinburg, who will support the findings by mathematical modelling. Altogether, the outcomes of this project will yield fundamental knowledge about regulation of microtubule cytoskeleton, which is relevant to human biology and disease.

References:

  • Gomez, J. M., Chumakova L., Bulgakova N. A., & Brown, N. H. (2016). Microtubule organization is determined by the shape of epithelial cells. Nature Communications, 7, 13172 doi: 10.1038/ncomms13172
  • Bulgakova, N. A., Grigoriev, I., Yap, A. S., Akhmanova, A., & Brown, N. H. (2013). Dynamic microtubules produce an asymmetric E-cadherin-Bazooka complex to maintain segment boundaries. The Journal of Cell Biology, 201(6), 887–901. http://doi.org/10.1083/jcb.201211159
  • Preciado López, M., Huber, F., Grigoriev, I., Steinmetz, M. O., Akhmanova, A., Koenderink, G. H., & Dogterom, M. (2014). Actin-microtubule coordination at growing microtubule ends. Nature Communications, 5, 4778. http://doi.org/10.1038/ncomms5778

Keywords: Cell Biology / Development, Molecular Biology

For informal enquiries about this project, please contact:

JAK/STAT singling and the regulation of the cytoskeleton - Dr Martin Zeidler

Supervisors:

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

We have previously used the excellent developmental genetics of the Drosophila model organism to investigate the relationship between the JAK/STAT signal transduction pathway and the behaviour of fly blood cells (termed haemocytes) (Bausek & Zeidler 2013, Bina et al 2010). In addition, other researchers have shown that the JAK/STAT pathway is required to rearrange the cytoskeleton of epithelial pole cells to a mesenchymal morphology (EMT) in the developing fly ovary (Silver & Montel 2001). Simultaneously, other studies also suggest a role for human STAT3 in regulating the effectors of EMT in cancers. Taken together, all these studies suggest that the JAK/STAT pathway plays a key role in the regulation of the cytoskeletal changes involved in cell shape changes.

Using technologies developed in Evans lab, we have now have tools available with which to examine the dynamic movements of the haemocyte cytoskeleton in vivo. Using these tools, the responses of cells to stimuli such as nearby wounding or apoptotic cells can be readily examined.

In this project the successful candidate will examine the roles of JAK/STAT pathway signalling and its target genes in vivo. This work will have a particular focus on the haemocytes associated with the third instar larval imaginal discs. In particular, apoptosis of imaginal disc cells requires haemocytes to alter their morphology so as to invade these tissues to remove apoptotic corpses. We now seek to characterise the behaviour of larval haemocytes in response to apoptotic imaginal cells.

Once this ‘wild type’ behaviour has been characterised you will investigate the effect of loss- and gain-of-function mutations in the JAK/STAT pathway (and pathway target genes) on this process and how these genetic manipulations change cellular function and cytoplasmic morphology.

Ultimately this project aims to cast light on the mechanisms underpinning the cellular changes that occur in cancer metastasis.

References:

  • Bausek N, Zeidler MP (2014)“Gα73Β is a downstream effector of JAK/STAT signaling and a regulator of Rho1 in Drosophila hematopoiesis” J Cell Sci 127(1) 101-110
  • Bina S, Wright VM, Fisher KH, Milo M, Zeidler MP. (2010) “Transcriptional targets of
  • Drosophila JAK/STAT pathway signalling as effectors of haematopoietic tumour
  • Formation” EMBO Rep. 11(3) 201-7
  • Silver DL, Montell DJ. Paracrine signaling through the JAK/STAT pathway activates invasive behavior of ovarian epithelial cells in Drosophila. Cell. 2001;107(7):831–41.
  • Wendt, MK, Balanis N, Carlin CR, Schiemann WP. STAT3 and epithelial–mesenchymal transitions in carcinomas JAK-STAT  2014 3(2): e28975

Keywords: Cell Biology / Development, Genetics

For informal enquiries about this project, please contact:

Generating distinct actin networks in a single cytoplasm: Structural studies to determine the impact of actin nucleators on filament structure - Professor Kathryn Ayscough

Supervisors:

This project is directly funded by a White Rose BBSRC DTP studentship.  European/UK students only – find out more on our funding webpage.

A fundamental understanding of how a cell responds to its environment, in order to drive changes to cell physiology, is critical if we are to make relevant and appropriate interventions in the context of disease states. Many years of research have demonstrated that the actin cytoskeleton is a focal point of cell regulation, however there are still large gaps in our understanding of mechanisms governing early steps in actin filament formation and how distinct filament organizations can co-exist in a single cytoplasm. This project will combine molecular biology and biochemistry with advanced electron cryomicroscopy (CryoEM) approaches to gain new structural insights into the process of actin nucleation. It will be supported by world-class labs with expertise in actin biochemistry and cell biology (Ayscough) and in structural analysis of macromolecular complexes using CryoEM (Bullough).
The overall hypothesis that will be tested in this project is that distinct nucleators impose structural constraints facilitating binding of distinct protein subsets to specific filaments.

The Ayscough lab has identified a novel actin filament nucleation and elongation activity in the yeast WASP homologue Las17.  The overall aim of this project is to understand how this nucleation function works and how it interfaces with other nucleating systems in the cell. 

Objectives:

  1. To undertake electron microscopy and reconstruction approaches to determine whether filaments generated in the presence of the WASP family nucleator differ structurally from those generated through other nucleating conditions. 
  2. Use mutagenesis to define key properties required for nucleating and elongating function and determine whether, and how mutations in the nucleator affect F-actin structure.
  3. To investigate how the presence of SH3 domain proteins influence the actin structures generated by the nucleating activity. 

Novelty. The Ayscough lab is pioneering studies in actin nucleation. This project aims to exploit findings emanating from our BBSRC grant and to initiate studies in a more structural direction to generate a greater depth of understanding of current biochemical and cellular findings.

References:

  1. Allwood EG, Tyler JJ, Urbanek AN, Smaczynska-de Rooij II, Ayscough KR. Elucidating Key Motifs Required for Arp2/3-Dependent and Independent Actin Nucleation by Las17/WASP (2016). PLOSOne 11(9):e0163177
  2. Palmer SE, Smaczynska-de Rooij II, Marklew CJ, Allwood EG, Mishra R, Johnson S, Goldberg MW, Ayscough KR. A Dynamin-Actin Interaction Is Required for Vesicle Scission during Endocytosis in Yeast. (2015) Curr Biol. 25:868-78.
  3. Urbanek AN, Smith AP, Allwood EG, Booth WI, Ayscough KR. A novel actin binding motif in Las17/WASP nucleates actin filaments independently of Arp2/3. (2013) Curr Biol 23: 196-203
  4. Jiang S, Wan Q, Krajcikova D, Tang J, Tzokov SB, Barak I, Bullough PA. (2015). Diverse supramolecular structures formed by self-assembling proteins of the Bacillus subtilis spore coat. Mol Microbiol. 97(2):347-59.

Keywords: Biochemistry, Molecular Biology, Structural Biology, Macromolecular Chemistry, Data Analysis

For informal enquiries about this project, please contact:

Bacterial-associated oncogenesis: Investigating the typhoid toxin of the persistent intracellular pathogen Salmonella Typhi in oncogenesis - Dr Daniel Humphreys

Supervisors:

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

The research group is funded by an MRC New Investigator Research Grant and a Department of Biomedical Science Start-Up Fund.

Background

The human intracellular bacterial pathogen Salmonella enterica serovar Typhi (S.Typhi) causes systemic typhoid fever resulting in 27 million cases of disease and 200,000 deaths each year. S.Typhi is a stealth pathogen and a hallmark feature is its ability to also establish persistent chronic infections in the gall bladder where the pathogen is associated with causing cancer. Oncogenesis is exacerbated in humans carrying gene mutations that predispose individuals to gall bladder cancer. This is relevant to public health, as a link between infection and tumour development may result in personalised therapeutic protocols.

How S.Typhi persists in host cells and promotes oncogenesis is not known. S.Typhi initiates infections by injecting virulence proteins into mammalian host cells to direct uptake and replication within intracellular Salmonella-containing vacuoles (SCVs). From its intracellular niche, S.Typhi secretes the typhoid toxin into the extracellular milieu where it enters target host cells. Once inside the cell the toxin traffics to the nucleus where it reprogrammes cell cycle progression, causes DNA damage and manipulates cellular signalling through nuclease and ADP-ribosylase activities. Recent evidence shows that typhoid toxin mediates establishment of persistent infections in animal hosts, and persistence underlies S.Typhi-associated tumour development.
Project

The mechanisms by which typhoid toxin manipulates cells to drive persistent infections are not understood, and the significance of the toxin to S.Typhi’s stealth virulence strategy and bacterial-associated oncogenesis has not been addressed. The PhD project will investigate these research questions using purified toxin derivatives and infection models in combination with the latest advances in molecular cell biology, fluorescence microscopy, and automated high-throughput screening technologies.

The student will join the laboratory of Dr. Daniel Humphreys (first supervisor) at the Department of Biomedical Science (BMS), University of Sheffield. The project will establish assays to study mammalian cells intoxicated with typhoid toxin using molecular, biochemical, genetic and fluorescence imaging approaches. In particular, how the toxin perturbs nuclear functions in cells predisposed to oncogenesis during S.Typhi infection will be examined using high resolution using structural resolution microscopy at the Wolfson Light Microscopy Suite at the BMS.
Having established the project foundations an RNAi screen will be developed with Dr. Steve Brown (third supervisor) at the RNAi Screening Facility, University of Sheffield.

The PhD student will be trained to use robotics and automated microscope systems to perform high-throughput RNAi screens aimed at identifying host genes required for cellular manipulation by the typhoid toxin.

Finally, the student will use Salmonella infection models, intoxication and cell transformation assays to investigate the significance of genes identified in the screen on pathogen persistence and the generation of bacterial-associated oncogenic phenotypes. These project components will be developed in the laboratories of the first supervisor and the second supervisor, Dr. Anjam Kahn, at the University of Newcastle.

To combat typhoid fever we need to understand the hijack mechanisms employed by S.Typhi to cause disease. By bringing together researchers from the University of Sheffield and the University of Newcastle, three areas of expertise will be combined to provide excellent scientific training and exposure to a broad range of techniques in a project of enormous biomedical significance. 

References

  1. Humphreys D*. Singh V, Koronakis V*. (2016). Inhibition of WAVE Regulatory Complex activation by a bacterial virulence effector counteracts pathogen phagocytosis. Cell Reports. 17(3):697-707. PMID: 27732847.
  2. Humphreys D, Davidson AC, Hume PJ, Makin LE, Koronakis V. (2013) Arf6 coordinates actin assembly through the WAVE complex, a mechanism usurped by Salmonella to invade host cells. Proceedings of the National Academy of Sciences of the United States of America. 110(42):16880-16885. PMID: 24085844
  3. Humphreys D, Davidson AC, Hume PJ, Koronakis V. (2012) Salmonella SopE and host GEF ARNO cooperate to recruit and activate WAVE to trigger bacterial invasion. Cell Host & Microbe 11, 129-39. PMID: 22341462
  4. Smith, K., Humphreys, D., Hume, P.J., and Koronakis, V. (2010) Enteropathogenic Escherichia coli recruits the cellular inositol phosphatase SHIP2 to regulate actin-pedestal formation.
    Cell Host & Microbe 7, 13-24. PMID: 20114025

Keywords: Biochemistry, Cancer / Oncology, Cell Biology / Development, Genetics, Microbiology, Molecular Biology, Pathology

For informal enquiries about this project, please contact:

Targeting dystroglycan to the nucleus in muscular dystrophy and cancer - Professor Steve Winder

Supervisor: Professor Steve Winder

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Open to self funded students as well as possible BMS funded.

Background

Dystroglycan is an essential cell adhesion receptor required for early embryonic development. Genetic loss of function gives rise to severe muscular dystrophies with neuronal involvement. Post-translational loss of function also occurs in Duchenne muscular dystrophy and in some cancers. This includes phosphorylation, proteolysis and ubiquitination. Moreover some proteolytic fragments of dystroglycan are targeted to the nucleus where they have effects on transcription. As part of our analysis of dystroglycan post-translational modifications, we identified a lipid modification - palmitoylation of a conserved cysteine residue that could act to anchor bioactive dystroglycan fragments to the membrane, both at the cell surface and in the nucleus.

Aims:

To examine the function of dystroglycan palmitoylation in cellular targeting and in cellular phenotypes associated with muscular dystrophy and cancer.

Techniques:

PCR-based site-directed mutagenesis, and cloning of dystroglycan mutants. Overexpression of dystroglycan mutants in tissue culture cells. Analysis of subcellular distribution of dystroglycan by quantitative immunofluorescence microscopy and cell fractionation. In vitro assays of cell invasion and metastatic growth.

References:

  • Leocadio D, Mitchell A, Winder SJ.
    γ-Secretase Dependent Nuclear Targeting of Dystroglycan.
    J Cell Biochem. 2016 Mar 18. doi: 10.1002/jcb.25537
  • Martínez-Vieyra IA, Vásquez-Limeta A, González-Ramírez R, Morales-Lázaro SL, Mondragón M, Mondragón R, Ortega A, Winder SJ, Cisneros B.
    A role for β-dystroglycan in the organization and structure of the nucleus in myoblasts.
    Biochim Biophys Acta. 2013 1833:698-711
  • G. Mathew, A.Mitchell, J.M.Down, F.C.Hamdy, C.Eaton, D.J.Rosario, S.S.Cross & S.J.Winder (2013)
    Nuclear targeting of dystroglycan promotes the expression of androgen regulated transcription factors in prostate cancer.
    Sci. Rep. 3, 2792.

Keywords: Biochemistry, Cancer / Oncology, Cell Biology / Development, Molecular Biology

For informal enquiries about this project, please contact:

An integrated molecular, cell and proteomic analysis of activity-dependent signalling in neurons - Dr Mark Collins

Supervisor: Dr Mark Collins

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Recent proteomic investigations have identified several thousand proteins in biochemically purified synapses and have uncovered multiprotein complexes essential for synapse function. Through these experiments, we have found that many of these synaptic genes are associated with over 100 neurological diseases, highlighting the need for a better a molecular understanding of synapses.

Many genes/proteins have been implicated in different aspects of synaptic plasticity in single gene/low-throughput studies. However, few studies have taken unbiased approaches to measure global synaptic responses at the protein level and therefore it is not clear what the relative significance of a change in expression or phosphorylation of a single protein in the context of the synapse proteome as a whole. The global analysis of early to late signalling events in synaptic stimulation time-course experiments will also add an important temporal dimension to our understanding of signal transduction and processing at synapses.

Furthermore, the extent and importance of receptor-mediated signalling pathway cross-talk is poorly characterised; the molecular dissection of post-translational modification cross-talk at the synapse will be important to understand how signal integration occurs through complex signalling networks. The use of mass spectrometry-based proteomics has great potential to further our understanding of synaptic signalling pathways in an unbiased, quantitative and temporal manner.

This interdisciplinary project will utilise a range of experimental approaches including cell culture, molecular and cell biology, biochemistry and state-of-the-art quantitative mass spectrometry. The student will be given in-depth training in all of these methods and will benefit from collaborations with other groups within the department.

The aims of the project include the following.

  1. Identify protein expression and post-translational modification (e.g. phosphorylation, ubiquitination and palmitoylation) changes that occur after established stimulation paradigms in neuronal cell culture.
  2. Examine temporal features and cross talk of signalling pathways in time course experiments
  3. Characterise the function of activity dependent post-translational modifications on selected disease-relevant genes

References:

  • Coba MP, Pocklington AJ, Collins MO, Kopanitsa MV, Uren RT, Swamy S, Croning MD, Choudhary JS, Grant SG. (2009).
    Neurotransmitters drive combinatorial multistate postsynaptic density networks.
    Science Signaling. Apr 28;2(68):ra19.
  • Ebert DH, Greenberg ME. (2013).
    Activity-dependent neuronal signalling and autism spectrum disorder.
    Nature. Jan 17;493(7432):327-37
  • Cox J, Mann M. (2011).
    Quantitative, high-resolution proteomics for data-driven systems biology.
    Annual Review of Biochemistry 2011;80:273-99.

Keywords: Biochemistry, Cell Biology / Development, Neuroscience/Neurology

For informal enquiries about this project, please contact:

The Role of cilia in processing of visual information - Dr Anton Nikolaev

Supervisors:

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Neurons of the vertebrate central nervous system, including these in the hippocampus and the cerebral cortex, are ciliated.  The function of these neuronal cilia remains, however, a mystery.  It is currently believed that central nervous system cilia may contribute to higher brain functions, such as memory and their malfunction may lead to psychiatric disorders, including schizophrenia and autism.

The proposed project will investigate the role of cilia in the processing of visual information in the retina using the zebrafish model. A combination of state-of-the-art techniques, including molecular genetics and 2-photon imaging of neuronal activity, will be used to study how the activity of visual neurons is affected by mutation causing abnormal ciliogenesis.

Keywords: Molecular Biology, Neuroscience/Neurology

For informal enquiries about this project, please contact:

Understanding how memory is recorded in the brain - Dr Anton Nikolaev

Supervisor:

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Understanding how memory is stored in the brain is one of the ultimate goals of neuroscience. The cellular basis of memory formation is a change in the synaptic strength during two processes: long term potentiation (LTP) and depression (LTD).  Molecular details of both LTP and LTD are well understood  (reviewed in Citry and Malenka, 2008) but exactly how changes in synaptic strength lead to memory formation is unclear. For example, it is unknown whether memory is stored in a large population of synapses or in changes of individual synapses.

It is also unclear how many bits (or perhaps bytes or even kilobytes?) of information is stored in an individual synapse and how synaptic plasticity is affected during multiple neurological disorders.  To investigate these kinds of problems requires in vivo imaging using optical reporters of synaptic plasticity in individual synapses.  One such reporter, AMPAR-pHluorin, reports trafficking of AMPA receptors during long-term potentiation. It has been used to study molecular details of LTP and LTD in cultured neurons (Ashby et al., 2004), while understanding how LTP and LTD encode memory needs to be performed on behaving animals.

The proposed project aims at using of AMPAR-pHluorin in vivo to understand how visual memory is stored in the zebrafish brain. The zebrafish brain is relatively simple, yet able to memorise a large number of different shapes. The project will utilise state-of-the-art imaging and molecular biology methods to answer the questions outlined above.

Keywords: Neuroscience/Neurology

For informal enquiries about this project, please contact:

Investigating perturbed signalling pathways underlying Motor Neurone Disease - Dr Mark Collins

Supervisor: Dr Mark Collins

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Motor Neurone Disease or Amyotrophic lateral sclerosis (ALS) is a disorder that results in fatal paralysis within a few years of symptom onset. Recently, a number of large exome sequencing studies of ALS patients have independently identified several loss of function mutations in the kinase, Tbk1. Repeat expansion in C9ORF72 is a major cause of ALS and results in the formation of aggregates that sequester RNA-binding proteins. Tbk1 phosphorylates optineurin and p62, cargo receptors for recruiting ubiquitinated proteins to the autophagosome for destruction. All three proteins are involved in autophagy and are relevant to the clearance of aggregates that result from C9ORF72 repeat expansion.

This project aims to understand how Tbk1 is involved in disease development and to identify potential targets for therapeutic intervention. This project will exploit a proximity labelling strategy to determine which signaling pathways downstream of Tbk1 are relevant to the formation and accumulation of protein aggregates characteristic of ALS.

This project will utilise a range of experimental approaches including molecular biology (cloning, mutagenesis), cell culture (including patient derived cell lines), protein biochemistry (western blotting, immunoprecipitation, kinase assays), imaging (immunofluorescence/confocal microscopy and state-of-the-art proteomic technologies (high resolution Orbitrap Mass Spectrometry).

References

  • Haploinsufficiency of TBK1 causes familial ALS and fronto-temporal dementia.
    Freischmidt A et al.
    Nat Neurosci. 2015 May;18(5):631-6.
  • Exome sequencing in amyotrophic lateral sclerosis identifies risk genes and pathways.
    Cirulli ET et al.
    Science. 2015 Mar 27;347(6229):1436-41.
  • Filling the Void: Proximity-Based Labeling of Proteins in Living Cells.
    Kim DI, Roux KJ.
    Trends Cell Biol. 2016 Sep 22. pii: S0962-8924(16)30134-9.
  • Decoding signalling networks by mass spectrometry-based proteomics.
    Choudhary C, Mann M.
    Nat Rev Mol Cell Biol. 2010 Jun;11(6):427-39.

Keywords: Biochemistry, Cell Biology / Development, Neuroscience/Neurology

For informal enquiries about this project, please contact:

How do post-translational modifications control the physiology of the human pathogen Campylobacter jejuni? - Dr Mark Collins

Supervisors:

This project is fully funded by a four-year White Rose DTP studentship in Mechanistic Biology from the BBSRC. Eligibility: UK/EU citizens only. EU citizens must have lived in the UK for at least three years to be eligible for full support. – find out more on our funding webpage.

BBSRC guide to studentship eligibility

Campylobacter jejuni is the commonest bacterial food-borne pathogen, responsible for an estimated 700,000 cases p.a. of gastroenteritis in the UK alone, mainly from undercooked chicken. The BBSRC priority strategy for eradicating this pathogen from the food chain demands greater understanding of its biology to identify new intervention targets. One important aspect of its metabolism that is poorly understood is the control of protein activity by post-translational modifications (PTMs).

We know that many essential proteins in bacteria are controlled by covalent modifications to their structure. This project seeks to understand the role of these modifications in C. jejuni, particularly phosphorylation, by comparing wild-type and various mutant strains, using state-of-the-art proteomics analyses employing high resolution mass spectrometry and combined with biochemical analysis of protein function. The student will be trained in molecular microbiology including bacterial physiology, mutant construction and global protein expression analysis, and will apply cutting-edge proteomic techniques to an important human pathogen.

This exciting new project will provide an excellent training in a wide range of techniques and has arisen as a result of collaboration between a molecular microbiologist (Dave Kelly) and an expert in post-translational modifications and proteomics (Mark Collins). We seek enthusiastic students who wish to join a team with an established reputation in Campylobacter biology and who wish to become expert in pathogen microbiology, gene cloning and mutagenesis, proteomics and mass spectroscopy as well as protein purification and biochemistry.

Keywords: Biochemistry, Microbiology, Molecular Biology

For informal enquiries about this project, please contact:

New therapeutics for motor neuron diseases - Professor Bazbek Davletov

Supervisor: Professor Bazbek Davletov

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

Motor neuron diseases such as Parkinson’s and Amyotrophic Lateral Sclerosis (ALS) often lead to autonomic dysfunctions including difficulty swallowing. Dysphagia (swallowing difficulties) is a major risk factor in motor neuron diseases because of lung infections and choking due to saliva production.

Currently, botulinum neurotoxin provides the most reliable, long-lasting partial reduction in salivation. The major obstacle in the use of botulinum neurotoxin stems from its dangerous paralysing properties restricting its application in patients which are already suffering from muscle weakness. We recently developed several non-paralysing botulinum molecules which we need to evaluate in relevant rodent models.

The successful PhD applicant will learn the production of botulinum-based drugs, develop quantitative method for measuring saliva production and investigate the novel therapeutics in rodents. The successful completion of this project will open new avenues for the treatment of motor neuron disease patients ultimately improving the quality of life for people with motor neuron diseases throughout the world.

References

  • Mangione AS, Obara I, Maiarú M, Geranton SM, Tassorelli C, Ferrari E, Leese C, Davletov B, Hunt SP.
    ‘Non-paralytic botulinum molecules for the control of pain.
    Pain 157:1045-55.
  • Ferrari E, Gu C, Niranjan D, Restani L, Rasetti-Escargueil C, Obara I, Geranton SM, Arsenault J, Goetze TA, Harper CB, Nguyen TH, Maywood E, O'Brien J, Schiavo G, Wheeler DW, Meunier F, Hastings M, Edwardson JM, Sesardic D, Caleo M, Hunt SP & Davletov B
    ‘A synthetic self-assembling clostridial chimera for modulation of sensory functions’
    Bioconjug Chem 24, 1750−1759.
  • Arsenault J, Ferrari E, Niranjan D, Cuijpers SAG, Gu C, Vallis Y, O'Brien J & Davletov B
    ‘Stapling of the botulinum type A protease to growth factors and neuropeptides allows selective targeting of neuroendocrine cells’
    J Neurochem 126, 223-33.

Keywords: Biochemistry, Biomedical Engineering, Biotechnology, Medical/Clinical Science, Molecular Biology, Neuroscience/Neurology, Pharmacology, Psychology & Psychiatry, Veterinary Medicine

For informal enquiries about this project, please contact:

Rme-6 as an integrator of endocytosis and signalling in cancer cells - Professor Elizabeth Smythe

Supervisor: Professor Elizabeth Smythe

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

The endocytic pathway regulates intracellular signalling in a variety of ways and when there are defects in this cross-talk, this can give rise to a variety of diseases including cancer. Rab5 is a major regulator of the early endocytic pathway, which cycles between an inactive, cytoplasmic GDP form and an active membrane-associated GTP form. Rab5 guanine nucleotide exchange factors (GEFs) catalyse the conversion of rab5 into its active form where it interacts with a variety of effectors which allow it to perform many different cellular functions, including signalling (Stenmark, 2009).

Our lab is particularly interested in Rme-6 which is a rab5 GEF that integrates signalling and trafficking of receptor tyrosine kinases such as EGFR and Tie2 by modulating flux through the endocytic pathway. Using SILAC mass spectrometry we have identified the interactome of Rme-6 in HeLa cells following EGF stimulation and have identified a number of novel binding partners.

The aim of this PhD project will  be to extend our current understanding of Rme-6 in the integration of endocytosis and signalling through examining the role of these novel binding partners of Rme-6 that interact in a cargo specific manner.

The project will utilise a variety of molecular cell biology approaches, including CRISPr technology to knockout genes of interest as well as knocking in tagged wild-type and mutant proteins, endocytic and signalling assays as well as high resolution light microscopy in fixed and living cells. There will be particular emphasis on the role of Rme-6 in cancer cells.

References

Stenmark, H.
Rab GTPases as coordinators of vesicle traffic.
Nat Rev Mol Cell Biol. 2009 10:513-25.

Semerdjieva S, Shortt B, Maxwell E, Singh S, Fonarev P, Hansen J, Schiavo G, Grant BD, Smythe E.
Coordinated regulation of AP2 uncoating from clathrin-coated vesicles by rab5 and hRME-6.
J Cell Biol. 2008 183:499-511.

Keywords: Cell Biology / Development

For informal enquiries about this project, please contact:

Structural analysis of Rme-6, a rab5GEF that integrates endocytosis and signalling - Professor Elizabeth Smythe

Supervisors:

This project is eligible for a department scholarship. These scholarships are awarded on a competitive basis – find out more on our funding webpage.

The ras family of small molecular weight GTPases act as molecular switches. In their active GTP conformation, they interact with a variety of effectors to perform a range of physiological functions. Conversion to the GTP conformation is mediated by guanine nucleotide exchange factors (GEFs) while GTPases are inactivated by GTP hydrolysis, facilitated by GTPase activating proteins (GAPs). Ras is the founding member of this family and has many roles in intracellular signalling and mutations in ras can result in cancer.

The rab family of small GTPases regulates many aspects of membrane trafficking and rab5 is considered to be a master regulator of the early endocytic pathway (Stenmark, 2009). Rme-6 is a multidomain protein containing an N-terminal rasGAP domain and a C-terminal rab5 GEF domain connected by a flexible linker. We have evidence that Rme-6 modulates endocytic flux of EGFR resulting in modulation of its downstream signalling by acting as a signalling scaffold.

We have explored the structure of hRme-6 and our recent results have shown that hRme-6 exists as a higher order multimer in vivo and in vitro. Furthermore we have shown its GEF and GAP activities are autoinhibited and require specific spatial activation in cells (2). Recent studies have identified the hinge region as key to regulation of Rme-6 conformation and relief of autoinhibition. We can purify Rme-6 and negative staining and EM analysis have provided a preliminary structure.

The aim of this project will be to refine this structure using cryo-EM approaches. We are particularly existed to understand the role of the hinge region in effecting conformational changes in Rme-6.

References

Stenmark, H.
Rab GTPases as coordinators of vesicle traffic.
Nat Rev Mol Cell Biol. 2009 10:513-25.

Semerdjieva S, Shortt B, Maxwell E, Singh S, Fonarev P, Hansen J, Schiavo G, Grant BD, Smythe E.
Coordinated regulation of AP2 uncoating from clathrin-coated vesicles by rab5 and hRME-6.
J Cell Biol. 2008 183:499-511.

Keywords: Biochemistry, Cell Biology / Development, Structural Biology

For informal enquiries about this project, please contact: