Ciccia, A.; (2006) The MUS81 family of proteins. Doctoral thesis , University of London.

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Abstract

The faithful and complete replication of DNA is necessary for the maintenance of genome stability. The endonuclease MUS81 has recently been implicated in the repair of blocked forks during DNA replication. MUS81 is related to the nucleotide excision repair proteins XPF and ERCC1, due to the common ERCC4 nuclease domain that they share. Based on database searches for proteins containing the ERCC4 domain, we have identified four novel members of the MUS81 family. We named two of them EME1 and EME2, because of their similarity with S. pombe Eme1 protein. We showed that EME1 and EME2 interact with MUS81 and that MUS81/EME1 and MUS81/EME2 complexes are endonucleases that exhibit a high specificity for synthetic replication fork and 3'-flap structures in vitro. In particular, the MUS81/EME2 heterodimer is 10-fold more active than MUS81/EME1. Besides EME1 and EME2, we have identified two additional proteins of the MUS81 family, HEF and HIP. HEF, also referred to as FANC-M, is a 250 kDa protein that is associated with the genetic disorder of Fanconi Anemia. HIP (HEF Interacting Protein) is a novel 24 KDa protein interacting with HEF/FANC-M. We showed that HIP forms a complex with HEF/FANC-M both in vitro and in vivo and that it is part of the Fanconi Anemia core complex. HEF/FANC-M contains a DEAH helicase domain, which is required for translocase activity, and an ERCC4 nuclease domain. We showed that the ERCC4 nuclease domain of HEF/FANC-M is inactive, as suggested by sequence analysis. Based on the similarity with other members of the MUS81 family, we propose a role for the complex between HEF/FANC-M and HIP in recognising branched DNA structures, which could arise after DNA replication fork blockage. Therefore, HEF/FANC-M and HIP may be involved in targeting the Fanconi Anemia core complex to blocked replication forks.

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