Expression of E-coli RecA targeted to mitochondria of human cells

Mitochondrial DNA integrity is ensured by several nuclear-encoded proteins in vertebrates, and a number of mtDNA alterations in human diseases, includ ing deletions and duplications, have been suspected to result from errors i n the mitochondrial recombination pathway. However, the presence of the lat ter system is still a matter of controversy as RecA proteins display variou s functions in vitro. In Escherichia coli, RecA plays a central role in hom ologous recombination by pairing and transfer ring a single strand to a hom ologous duplex DNA. To address indirectly the issue of a mitochondrial reco mbination pathway in vivo, we have constructed a chimeric gene containing a n N terminal mitochondrial targeting sequence and the E. coli RecA gene. Ce lls were transfected by the recombinant plasmid, then tested for their mtDN A repair upon bleomycin treatment. We found an increased repair rate of the mitochondrial DNA in cells expressing RecA as compared to control cells. T hese results indicate that the transfected cells display an improved mtDNA repair replication pathway due to the exogeneous RecA, likely in synergy wi th an endogeneous rate-limiting mitochondrial recombination pathway. (C) 20 01 Elsevier Science B.V. All rights reserved.