University of Dundee

Experimental and theoretical analysis of a novel genome stability pathway

During the eukaryotic cell division cycle, the genome must be precisely duplicated with no sections left unreplicated and no sections replicated more than once. Failure to do this causes major genome abnormalities including deletion and amplifications, which can drive a range of pathologies such as cancer. We have recently provided evidence for a new pathway allowing cells with incompletely replicated genomes to undergo cell division, allowing daughter cells to complete replication. This PhD project will use a combination of experimental and theoretical approaches to explore this novel pathway.

Our recent work in humans cells shows that at the end of S phase there are frequently a few stretches of DNA that remain unreplicated. Cells with unreplicated DNA can enter mitosis and segregate the unreplicated DNA to daughter cells via structures called ultrafine anaphase bridges. In these daughter cells a protein called 53BP1 then binds to the unreplicated DNA.

The PhD project will explore the mechanisms that allow unreplicated DNA to be segregated to daughter cells. We will examine the role of DNA repair pathways in protecting unreplicated DNA from being degraded before cells enter mitosis. We will examine how cells solve the challenge of unwinding the two unreplicated template strands so that they can be segregated to daughter cells. We will also examine the role of 53BP1 in protecting these structures during G1.

The project will use a combination of experimental and theoretical approaches, and will build on a successful collaboration that has been established between Prof Blow (experimental) and Prof Newman (theoretical). Experimental work will involve cell biology approaches using human tissue culture cells (including immunofluorescence, FACS, siRNA, CRISPR/CAS) and biochemical approaches (immunoblotting, immunoprecipitation, recombinant protein work) using Xenopus (frog) egg extracts. There are also opportunities for theoretical work involve a range of mathematical and physical modelling approaches to analyse the consequences of fork stalling.

Newman, T.J., Mamun, M.A., Nieduszynski, CA. and Blow, J.J. (2013). Replisome stall events have shaped the distribution of replication origins in the genomes of yeasts. Nucleic Acids Res. 41, 9705-9718.

Moreno, A., Carrington, J.T., Albergante, L., Al Mamum, M., Haagensen, E.J., Komseli, E.-S., Gourgolis, V.G., Newman, T.J. and Blow, J.J. Unreplicated DNA remaining from unperturbed S phases passes through mitosis for resolution in daughter cells. Proc Natl Acad Sci USA, in press.

Al Mamum, M., Albergante, L., Moreno, A., Carrington, J.T., Blow, J.J. and Newman, T.J. Inevitability and containment of replication errors for eukaryotic genome lengths spanning Megabase to Gigabase. Proc Natl Acad Sci USA, in press.