Department of Animal and Plant Sciences projects

Projects:

A genome-wide association study to reveal the wider signalling network of mTor – a key pathway in ageing

Main Supervisor:

Dr Mirre Simons

m.simons@sheffield.ac.uk

Second Supervisor:

Laura Hartshorne (Technician)

Type of Project:

Lab/Bench Project - primarily working in a lab environment

Aims and Objectives:

The best-established drug to extend life is rapamycin. Reduced mTor (mechanistic target of rapamycin) signalling extends lifespan in model species. Many inputs and outputs of the mTor signalling pathway have been identified, however possible interactions and the specific contributions of different outputs in determining lifespan, and their connections to metabolism and diet, are unclear. Moreover, because mTor is also strongly involved in development, manipulations of this pathway are required in the adult stage only. Advanced available genetic tools in flies (Drosophila melanogaster) allow temporal manipulation of gene expression to do just that: knockdown mTor expression in adult flies.

From recent data we gathered in my lab we now know that conditional knockdown of mTor leads to a rapid reduction in egg laying. You will use this novel phenotype to perform a genetic screen (using genome wide-association) for regulators of the mTor network in an in vivo multicellular system. Flies are ideal for student projects as they allow rapid data generation, follow-ups to be designed by the student and you will be able to quickly gain independence in your lab work, and work with living animals (not ethical approvals required). You will be able to test identified candidates (using in vivo RNAi approaches) yourself in the timeframe to experimentally link the identified candidates to egg laying and also lifespan.

Your approach to identify novel components of the mTor network will be to cross the conditional knockout model of mTor (already available) to a panel of 200 inbred lines (also already available) of which we have whole genome information (work with F1 only). You will then count eggs laid on two different diets, with and without mTor knockdown. These phenotypic data, combined with the genomic data (statistics to do this are in place) will allow a genome-wide association study to link SNPs to phenotype. These SNPs will be investigated in bioinformatic databases and candidate genes identified through that process can be manipulated in vivo using RNAi.

Research Methodology:

You will learn the following techniques. We have direct and proven experience with all these techniques.

• Genome-Wide Association using the latest statistical tools
• Bioinformatic analysis of the candidate genes
• In vivo RNAi and qPCR follow-up where relevant
• Egg counting using available software
• Virgin collection for crosses
• Fly husbandry
• Planning large experiments and handling large datasets

Timeline:

Aug: lab induction

Sept-Dec: phenotyping of DGRP flies crossed with inducible knockdown of mTor fly (all available)

Jan: Analysis in GWAS framework

Feb-April: Follow ups using in vivo RNAi

May: Write-up & apply for conference presentation where relevant

Expected Outcome:       

• A genome wide association study of signalling components of the mTor network.
• Manipulation of candidate genes in the egg laying, diet and lifespan context to test for causality of the identified genes.
• Extension and integration of our current knowledge of the mTor signalling network

Additional Training:

We will train you in all the methodology requires using one-on-one tutoring. You will further use to learn R, a command line environment we use for all stats and a large amount of data processing. You will need to learn R, in my view, if you want to do anything remotely quantitative in life sciences. You will be expected to act as a full member of our group, give presentations on your data, and join in with discussions on the other topics in my group.

Ethical Approval:

Non-human tissue - no ethics approval required.