Fanconi anemia (FA) is a genetic disease with defects in development and hematopoiesis and propensity to cancer, indicating a vital and basic cell biology process at work. The hallmark of FA is genomic instability, evidenced bv gross chromosomal breakage and DNA alkylating agent hypersensitivity, which correlates with cancer susceptibility in general. Studies of FA are important in several ways. First, FA biology is involved across a spectrum of scientific disciplines. Including hematology oncology, and development. Second, since the known FA proteins are found only in mammalian cells and have no previously described protein domain, their study will yield the description of a novel pathway which promotes the maintenance of genomic stability. Third, work on other cancer susceptibility syndromes have proved to have wide applicability in science in general and cancer in particular, such as Li- Fraument syndrome (p53), ataxia telangiectasia (P13 kinase), and xeroderma pigmentosum (DNA nucleotide excision repair). Fourth, basic work on FA has already led to clinical use of reagents for diagnosis and genetic counseling, and gene therapy trials are currently underwav for treatment of FA. The focus of the grant is to determine the specific biochemical nature of the FA pathway. The emphasis is on the FA proteins, which are encoded by the FA genes, accounting for 5 of at least 7 complementation groups. The FA proteins bind together in a protein complex which forms and is nuclear in 6 of the 7 FA complementation groups, which indicates its importance in FA biology. Our hypothesis is that the FA nuclear protein complex functions as a multimeric complex which is regulated by postranslational phosphorylation. Specifically the purpose of this grant is to isolate the complex in order to purify additional binding proteins of the FA complex to lend an idea to the overall function of the FA complex. In my preliminary, studies I have isolated FA binding proteins whose interactions with FA proteins will be confirmed and whose hypothetical functions will be tested in the work proposed. In addition we will demonstrate how modifications of proteins and of the size of the FA protein complex regulates its function and localization. Our recent work has shown that the FancG protein becomes phosphorylated at mitosis and the entire complex exits condensed chromosomes. We will map the phosphorylation site, test for the kinase involved and assess the changes in complex size in response to multiple stimuli. Identification of new proteins and elucidation of FA pathway mechanisms promise to shed light on a novel area of cancer biology with the potential to provide direct clinical applicability.
