Professor Walter Marcotti
Professor of Sensory Neuroscience
Room: B1 221 Alfred Denny building
Brief career history
Awards and prizes
Auditory neuroscience and Deafness Sensory transduction Synaptic transmission
Sensory organs and the neural networks responsible for processing sensory information are supremely well adapted for detecting input from the external environment. Their challenge is to maximize sensitivity and fidelity over a wide dynamic range. The sensory receptors of the mammalian auditory system, the inner hair cells (IHCs), do this with unparalleled temporal precision (kHz range). We know little about the molecular and physiological mechanisms controlling the functional maturation of the auditory system or signal processing at the primary auditory synapses, the IHC ribbon synapses. Crucial to this work, is the need of near-physiological in vitro and the development of in vivo experimental models.
My laboratory is uniquely suited for this task because it is the only one in the world that routinely uses near-physiological conditions for in vitro mammalian cochlear physiology and performs in-vivo electrophysiology from the zebrafish. How biological systems orchestrate their development and how complex signals are processed by mature neuronal networks are major challenges in the quest to understand human biology and disease.
The auditory system provides an ideal model with which to address these questions, primarily because it involves a highly ordered array of a very small number of sensory cells with well-defined neuronal circuitry. It is also a key priority for human health because hearing loss affects more than 360 million people worldwide (WHO 2013), a number that will increase with the aging population.
Undergraduate and postgraduate taught modules
Dystroglycan function in mammalian auditory hair cells and neurons - Action on Hearing Loss Studentship
Action on Hearing Loss PhD Studentship (3 year fully funded PhD studentship (Uk and EU only)
Supervisors: Professor Steve Winder and Professor Walter Marcotti, Department of Biomedical Science
This PhD offers an exciting opportunity for a talented student to work on a multidisciplinary project in the areas of auditory physiology and molecular cell biology investigating the role of dystroglycan in mammalian hearing.
This proposal focuses on the function of a transmembrane protein called b-dystroglycan, which forms the core of dystroglycan complexes that enable cells to establish mechanical connections and communication pathways between their internal structures and the extracellular environment.
The inner ear interprets mechanical vibrations induced by sound. This requires the precise transduction of incoming auditory stimuli into an electrical signal that can be perceived by the brain. This sophisticated biological process occurs at the sensory hair cells. The hair cell mechanotransducer apparatus projects into a unique extracellular fluid compartment that is rich in potassium ions and that provides the ‘battery’ for the inner ear. Furthermore, hair cells transfer electrical information rapidly to their sensory nerves via specialised ‘ribbon’ synapses. All of these processes are expected to involve dystroglycan complexes, although published evidence suggests that b-dystroglycan is not present in the sensory organ. This is puzzling because there is clear evidence for its partner, a-dystroglycan, which is encoded by the same gene. We also know that defects in a number of other proteins linked to dystroglycan complexes are associated with hearing loss. One of these proteins is dystrophin, mutations of which is normally linked to muscular dystrophy.
Recently, we found that tyrosine phosphorylation of b-dystroglycan occurs at unusually high levels in the ear and that this explains why the protein was not previously detected. It changes the binding properties to other proteins, which is of general biological interest because dystroglycans are ubiquitously expressed yet they serve a wide range of different functions. In muscle, dystrophin binds to dystroglycan complexes and dissipates contraction forces to the extracellular connective tissue, thus protecting muscle membrane integrity. Mutations in dystrophin weaken these connections so that muscle cells become more easily damaged and degenerate, leading to muscular dystrophy. The strength of the attachment can be altered through the modification to b-dystroglycan that we observe in the ear and the mechanism has been proposed as a potential therapy to stabilise the muscle. Similar modifications to b-dystroglycan act as switches to alter association with other proteins in non-muscle cells, including nerves, and they are important for the organisation of signalling molecules at synaptic junctions and of molecules that regulate the flow of water and inorganic ions across fluid compartments. Finally, longer term changes in the function of dystroglycan complexes have been associated with aging. Thus phosphorylation of b-dystroglycan in the ear is likely to be critical for auditory function and for greater understanding of noise-induced and age-related hearing loss.
Outline of the research methods
Animal models: We have made a knock-in mouse (Dag1Y890F) with a phenylalanine substitution that prevents tyrosine phosphorylation of β-dystroglycan . This will allow us to focus on the specific function of this modification in the ear. We will characterise the model, using the structural, physiological and biochemical techniques listed below. To identify potential hearing loss linked to the inability to phosphorylate Y890 in b-dystroglycan we will perform in vivo electrophysiological recording (auditory brainstem responses, ABRs) from normal and Dag1Y890F mice at 1, 4, 8 and 12 months old at frequencies of 4, 8, 16 and 32kHz.
Cell lines and antibodies: We have a conditionally immortal, mouse otic neuronal cell line, US/VOT-N33  that expresses β-dystroglycan and eps8, which has been assayed both by oligonucleotide gene expression arrays and by immunolabeling. The cells also react strongly with antibodies to pY890 b-dystroglycan. Eps8 is known to be expressed in neurons and it is associated with synaptic plasticity and the morphology of dendritic spines . We have the hybridoma cell line that produces antibodies to non-phosphorylated β-dystroglycan (Mandag2) and our own polyclonal antibody to pY890 b-dystroglycan (Ab1709) with which we will be able to estimate relative expression of the two forms in tissue sections, organotypic whole-mounts of the organ of Corti and in western blots and immunoprecipitations. We will use antibodies to ribeye (ribbon synapse), eps8 and eps8l2 (hair bundles and neurons), and Kir4.1 and aqp4 (fluid regulation) to study the distribution of these proteins in Dag1Y890F mice.
Single cell electrophysiology - Patch clamp recording will carried out on hair cells as described previously . We will use acutely dissected tissue to record mechanoelectrical transducer current from hair cells in response to displacements of hair bundles with a fluid jet and to assess basolateral membrane properties such as ion currents and vesicle release at the ribbon synapses.
Microscopy – Confocal fluorescence microscopy will be used to localise proteins in cells, cryostat sections and organotypic cultures. Scanning and transmission electron microscopy will be used to study the hair bundles and the localisation and structure of ribbon synapses.
Biochemistry – We will validate the Eps8 SH3 domain interaction with β-dystroglycan using peptide SPOT arrays to define the precise binding site on dystroglycan and using reciprocal fusion protein pull down assays from otic epithelial cell lysates. The interaction will be further validated by immunoprecipitation, immunofluorescence localisation and proximity ligation in otic epithelial cells in vitro. We have used similar approaches to validate SH3 domain interactions between β-dystroglycan and other cytoskeletal proteins [5, 6].
 Miller G, Moore CJ, Terry R, et al. Preventing phosphorylation of dystroglycan ameliorates the dystrophic phenotype in mdx mouse. Hum Mol Genet 2012;21:4508-20.
For informal enquiries about this project, please contact:
Professor Steve Winder
Professor Walter Marcotti
- Johnson SL, Ceriani F, Houston O, Polishchuk R, Polishchuk E, Crispino G, Zorzi V, Mammano F & Marcotti W (2017) Connexin-Mediated Signaling in Nonsensory Cells Is Crucial for the Development of Sensory Inner Hair Cells in the Mouse Cochlea.. J Neurosci, 37(2), 258-268. View this article in WRRO
- Johnson SL, Olt J, Cho S, von Gersdorff H & Marcotti W (2017) The coupling between Ca2+ channels and the exocytotic Ca2+ sensor at hair cell ribbon synapses varies tonotopically along the mature cochlea.. Journal of Neuroscience. View this article in WRRO
- Olt J, Allen CE & Marcotti W (2016) In vivo physiological recording from the lateral line of juvenile zebrafish.. Journal of Physiology, 594(19), 5427-5438. View this article in WRRO
- Corns LF, Johnson SL, Kros CJ & Marcotti W (2016) Tmc1 Point Mutation Affects Ca2+ Sensitivity and Block by Dihydrostreptomycin of the Mechanoelectrical Transducer Current of Mouse Outer Hair Cells. Journal of Neuroscience, 36(2), 336-349.
- Corns LF, Johnson SL, Kros CJ & Marcotti W (2014) Calcium entry into stereocilia drives adaptation of the mechanoelectrical transducer current of mammalian cochlear hair cells. Proceedings of the National Academy of Sciences, 111(41), 14918-14923.
- Furness DN, Johnson SL, Manor U, Rüttiger L, Tocchetti A, Offenhauser N, Olt J, Goodyear RJ, Vijayakumar S, Dai Y, Hackney CM, Franz C, Di Fiore PP, Masetto S, Jones SM, Knipper M, Holley MC, Richardson GP, Kachar B & Marcotti W (2013) Progressive hearing loss and gradual deterioration of sensory hair bundles in the ears of mice lacking the actin-binding protein Eps8L2.. Proc Natl Acad Sci U S A, 110(34), 13898-13903.