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Title: Functional characterisation of mammalian GEMC1
Author: Piergiovanni, G. M. M.
ISNI:       0000 0004 5362 8109
Awarding Body: University College London (University of London)
Current Institution: University College London (University of London)
Date of Award: 2014
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Chromosomal DNA must be faithfully duplicated only once per cell cycle in order to ensure genome stability. At the end of mitosis and during G1 phase of the cell cycle, eukaryotic cells assemble pre-replicative complexes (pre-RCs) on multiple replication origins distributed along their chromosomes in a process called origin licensing. During S phase, licensed origins are activated, DNA is unwound and the replicative machinery is recruited triggering origin firing. Cyclin-dependent kinases activity (CDKs), ubiquitin-dependent protein proteolysis and licensing factors inhibitors cooperate to promote origins firing and prevent replication origins from being reused within the same cell cycle. Mis-regulation of origin licensing contributes to genetic instability and is commonly observed in a variety of cancers. GEMC1 is a recently identified CDK target involved in the control of DNA replication in Xenopus laevis. GEMC1 is highly conserved in vertebrate organisms but nothing is currently known about its biological role in mammalian cells. In this study, the first characterisation of mammalian GEMC1 is presented. GEMC1 is a nuclear protein that is highly regulated during the cell cycle by a series of post- translational modifications, which include CDK-dependent phosphorylation and ubiquitination. Similar to its Xenopus homologue, GEMC1 interacts with TopBP1 and Cdc45 in S phase and its depletion strongly impairs origins firing, leading to the accumulation of DNA double-strand breaks (DSBs) in replicating mouse and human cells. These results confirm that mammalian GEMC1 has a positive role in chromosomal DNA replication. GEMC1 also forms a complex with the licensing inhibitor Geminin, both in vivo and in vitro, through its coiled-coil domain. Strikingly, GEMC1 overexpression in human U2OS cells induces DNA re-replication, chromosomal breakages that cause a G2/M arrest and centrosome amplification. All together, these results suggest that GEMC1 is a powerful DNA replication promoting factor that functions by stimulating origin firing and by antagonising Geminin dependent inhibition of origin licensing in vertebrate cells. These data also indicate that regulation of GEMC1 protein levels is critical for the maintenance of genome stability.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available