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Function of Mte1/ZGRF1 in Homologous Recombination
Author(s) -
Durham Jesse
Publication year - 2020
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2020.34.s1.09324
Subject(s) - rad51 , homologous recombination , biology , mutant , dna repair , replication protein a , fanca , mutagenesis , genetics , fanconi anemia , microbiology and biotechnology , mutation , dna , dna binding protein , gene , transcription factor
Fanconi anemia (FA) is a human genetic disorder that causes organ defects, physical abnormalities, bone marrow failure, and increased risk of malignancy. These phenotypes result from an impaired response to DNA damage due to the dysfunction of a cluster of FA proteins. Among these, human FANCM (budding yeast ortholog, Mph1) functions in DNA replication fork (RF) repair and DNA double‐strand break (DSB) repair by homologous recombination (HR). In a recent study, Mte1 in Saccharomyces cerevisiae was found to protect the Rad51‐made D‐loop, essential for HR, from disassociation by Mph1 (Xue et al. 2016). Our preliminary data showed that Mte1 interacts with Rad51 in vitro. However, the biological role of this interaction remains unknown. The goal of this project is to examine the effects of 2 different binding site mutants of Mte1, where mutations are hypothesized to affect binding of Rad51. Quick change site directed mutagenesis will be used to create the 3 mutants by changing 2 different phenylalanine side groups to alanine at positions 156, 169, and 156/169 combined. The Rad51 interaction of each mutant will be evaluated via in vitro pull down assay. The D‐loop binding activity of each mutant will be evaluated via electrophoresis mobility shift assays (EMSA). Mutation effects on Rad51‐made D‐loop formations can direct further studies investigating proteins required for HR. Evidence suggest that human ZGRF1 is functionally related to Mte1 due to N‐terminus conservation between the two. However, little is known about the role and function that ZGRF1 plays in HR in humans. Support or Funding Information Research supported by NIH R21 #ES028792 awarded to XX. JD supported by NIH R25 Bridge to Doctorate Program (GM102783)