About the Project
Summary
Stem cell therapy has to potential to revolutionize the way we treat human disease. But success will rely upon developing our understanding of stem cell biology. Stem cells function both to produce differentiated cells and maintain a stem cell population. The Drosophila testis is an ideal model in which to study germ stem cells and their niche. Regulation of gene expression is vital in controlling the balance between self-renewal and differentiation. Recent experiments indicate that mRNA translation regulation plays roles in maintenance and differentiation.
This project will investigate the role of RNA-binding proteins in the regulation of mRNA translation events in the testis and their importance to germ stem cell function. The molecular mechanisms underlying this control of protein synthesis will be characterized through biochemical and structure analysis (eg. X-ray crystallography, with the potential to take advantage of the University’s recent £16 million investment in the Astbury Biostructure Laboratory). The extent of mRNA translational regulation with be assessed using Next Generation Sequencing methods and complemented with functional analysis by Drosophila genetics.
Background
STAR (signal transduction and activation of RNA) proteins are KH-domain-containing, RNA-binding proteins whose activity is regulated in response to signals and regulate RNA processes. One family member is Drosophila HOW, which is expressed in the brain, testes and muscles. Mutants exhibit slow heart rates, heart defects and male sterility. Orthologs have been shown to regulate mRNA translation eg. p53 mRNA in C.elegans. HOW is required for germ stem cell maintenance in Drosophila testes and binds to RNAs that regulate differentiation in testes.
Objectives
1) Identify mRNAs translationally regulated by HOW in testes
2) Characterise molecular mechanism of translational regulation by HOW
3) Understand role of HOW regulated translation events in germ cells
Novelty
Transciptome-wide studies of translation have only been possible in the last few years, with most determining regions of translation and translational efficiencies. Combining ribosome profiling with structural analysis and Drosophila genetics is an innovative strategy, to dissect the molecular mechanisms of translational control by a specific protein.
Timeliness
The advent of ribosome profiling recently allowed global assessments of translational regulation. Improvements to the method, including ours (Aspden eLife 2014) now allow translational regulation by specific proteins to be measured. The dogma has been that the majority of gene expression regulation occurs at transcription but recent work has revealed mRNA translation is also a key point of regulation, eg binding of proteins to mRNAs alters the ability of ribosomes to initiate translation. Structural studies of STAR-proteins are currently at the point (including Edwards group), where mechanistic insight into how binding affects RNA activity can be achieved.
Experimental Approach
-Next Generation Sequencing to study translational control (ribosome profiling)
-Biochemical structural methods to understand how RNA-binding occurs and how this affects the ribosome (X-ray crystallography)
-Drosophila genetic analysis to assess how molecular interactions affect germ stem cell biology and fly fertility.
Keywords:
RNA binding proteins, germ stem cells, mRNA translation, Next Generation Sequencing, structural biology, fertility.
The student will be co-supervised by Dr. Julie Aspden, Dr. Tom Edwards and Dr Amanda Bretman.
More information on Aspden Lab: http://www.fbs.leeds.ac.uk/staff/profile.php?tag=Aspden_J
Informal enquires: [Email Address Removed]
Funding Notes
4 year BBSRC studentship, under the White Rose Mechanistic Biology DTP. The successful applicant will receive fees and stipend (c.£14,057). The PhD will start in Oct 2016. Applicants should have, or be expecting to receive, a 2.1 Hons degree in a relevant subject. EU candidates must have been resident in the UK for 3 years in order to receive full support. There are 2 stages to the application process: http://www.fbs.leeds.ac.uk/postgraduate/phdopportunities.php
References
Aspden, J. L., Eyre-Walker, Y. C., Philips, R., Amin, U., Mumtaz, A. S., Brocard, M., Couso, J. P. Extensive translation of small ORFs revealed by Poly-Ribo-Seq. eLife 10.7554/eLife.03528 (2014).
Jenkins, H.T., Malkova, B., Edwards, T.A., Kinked β-strands mediate high-affinity recognition of mRNA targets by the germ-cell regulator DAZL. PNAS 108 18266-18271 (2011).
Ariza, A., Tanner, S.J., Walter, C.T., Dent, K.C., Shepherd, D.A., Wu, W., Matthews, S.V., Hiscox, J.A., Green, T.J., Luo, M., Elliott, R.M., Fooks, A.R., Ashcroft, A.E., Stonehouse, N.J., Ranson, N.A., Barr, J.N., Edwards, T.A., Nucleocapsid protein structures from orthobunyaviruses reveal insight into ribonucleoprotein architecture and RNA polymerization. Nucleic Acids Research 41 5912-5926 (2013).
Price, T. A., Lizé, A., Marcello, M., Bretman, A. Experience of mating rivals causes males to modulate sperm transfer in the fly Drosophila pseudoobscura. J Insect Physiol 58 1669-1675 (2012).
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