Identification and Characterization of Novel Factors Involved in Interstrand Cross-link Repair

The hallmark of Fanconi anemia (FA) cells is their hypersensitivity to DNA interstrand cross-linking (ICL) agents. It is known that all the FA proteins cooperate in a common pathway that is activated after treatment with ICL agents, but exactly how this pathway leads to DNA repair is still obscure. Our aim is to identify and characterize novel enzymes that may play a role in ICL repair and interact with FA proteins. In Drosophila melanogaster, the mus308 mutation leads to marked sensitivity to ICLs, thus resembling FA cells. The C-terminal portion of the Mus308 polypeptide encodes a DNA polymerase, while the N-terminal portion encodes a DNA helicase. We cloned three mus308 related genes: POLQ, HEL308 and POLN both from human and mouse. We purified recombinant proteins and assayed their activity "in vitro". We also analyzed their expression "in vivo", both in mouse and human tissues. We have some evidences suggesting that human POLN might be involved in ICL repair, thus possibly interacting with the FA pathway. In order to understand the roles of the Mus308 related genes and FA proteins in ICL repair, we studied the genetically amenable multicellular model system C. elegans, and characterized the phenotype of worms that do not express these genes. We will present our recent data that indicate a possible function of the Mus308 family downstream the FA pathway. Our goal is to find a connection between the FA signal transduction pathway and the enzymes that physically participate in the removal of ICLs.