Pepe, A; (2014) Functional Characterisation of Human MUS81 Complexes. Doctoral thesis (PhD), UCL (University College London).

Preview

Text Pepe_A_thesis.pdf Download (33MB) | Preview

Abstract

Human cells employ a variety of mechanisms to repair DNA damage and maintain genomic integrity. Structure-specific endonucleases play crucial roles in the repair of DNA lesions by removing secondary DNA structures that can lead to chromosome mis-segregation, aneuploidy and cancer. MUS81 is the catalytic subunit of two human structure-specific endonucleases, MUS81-EME1 and MUS81-EME2. MUS81 nuclease has been shown to function in the resolution of homologous recombination (HR) intermediates, in the repair of stalled replication forks (RFs) and in the maintenance of telomere length in alternative lengthening of telomeres (ALT)-positive cells. However, it is unknown whether the two MUS81 complexes differ in their biological functions. To address this question, we carried out in vivo and in vitro studies to determine the roles of MUS81-EME1 and MUS81-EME2 in human cells. We found that EME2 interacts with MUS81 preferentially during the S-phase of the cell cycle, when MUS81 is important for the repair of stalled RFs. Cells treated with HU and depleted of MUS81 or EME2, but not of EME1, showed high levels of chromosomal aberrations, suggesting that the MUS81-EME2 complex is important for the maintenance of genomic stability following HU exposure. Also, depletion of EME2 or MUS81, but not of EME1, resulted in telomere loss, decreased rate of telomere sister chromatid exchanges (T-SCEs) and increased telomere fragility in ALT-positive cells. Consistent with the in vivo functional differences, we found that purified MUS81 complexes have different DNA substrate specificities in vitro, with the activity of MUS81-EME2 being 10-fold greater than that of MUS81-EME1. Together, these results indicate that MUS81-EME1 and MUS81-EME2 have different biochemical properties and distinct, non-overlapping functions, with MUS81-EME2 being important in the repair of stalled RFs and in telomere maintenance of ALT cells, whereas MUS81-EME1 plays an important role in the resolution of HJs in the context of HR-mediated DNA repair.

Download activity - last month

Export as JSONXMLCSV

Download activity - last 12 months

Export as JSONXMLCSV

Downloads by country - last 12 months

Export as JSONXMLCSV

Archive Staff Only

View Item