The FANCJ/MutLα interaction is required for correction of the cross-link response in FA-J cells

Abstract

FANCJ also called BACH1/BRIP1 was first linked to hereditary breast cancer through its direct interaction with BRCA1. FANCJ was also recently identified as a Fanconi anemia (FA) gene product, establishing FANCJ as an essential tumor suppressor. Similar to other FA cells, FANCJ-null (FA-J) cells accumulate 4N DNA content in response to DNA interstrand crosslinks (ICLs). This accumulation is corrected by reintroduction of wild-type FANCJ. Here, we show that FANCJ interacts with the mismatch repair complex MutLa, composed of PMS2 and MLH1. Specifically, FANCJ directly interacts with MLH1 independent of BRCA1, through its helicase domain. Genetic studies reveal that FANCJ helicase activity and MLH1 binding, but not BRCA1 binding, are essential to correct the FA-J cells' ICL-induced 4N DNA accumulation and sensitivity to ICLs. These results suggest that the FANCJ/MutLa interaction, but not FANCJ/BRCA1 interaction, is essential for establishment of a normal ICL-induced response. The functional role of the FANCJ/MutLa complex demonstrates a novel link between FA and MMR, and predicts a broader role for FANCJ in DNA damage signaling independent of BRCA1.

A novel protein, MAPO1, that functions in apoptosis triggered by O6-methylguanine mispair in DNA

“…YT103 and YT102M cells displayed increased resistance to MMS in comparison to YT102, as reported earlier (Kaina et al, 1991;Glaab et al, 1998). YT102 cells were quite resistant to ACNU as expected (Peng et al, 2007; -methylguanine-DNA methyltransferase (MGMT) and mismatch repair (MMR) proteins in YT102, YT103, KH101 and YT102M cell lines. The MGMT and MMR proteins were detected with immunoblotting using specific antibodies.…”

“…The MMR protein-dependent release of cytochrome c from mitochondria and activation of caspases, which are hallmarks for apoptosis, also occur in the case of O 6 -methylguanine-induced apoptosis (Ochs and Kaina, 2000;Takagi et al, 2003;Hickman and Samson, 2004). As apoptosis caused by bulky DNA lesions that block DNA replication occurs even in cells deficient in MMR proteins (Peng et al, 2007;Sanada et al, 2007), the apoptosis pathway induced by O 6 -methylguanine, which does not block DNA replication, appears to be distinct from the former. It is highly likely that other proteins are also involved in the pathway.…”

Insight into the Roles of Helicase Motif Ia by Characterizing Fanconi Anemia Group J Protein (FANCJ) Patient Mutations

“…Because Western blot analysis demonstrated that GFP-tagged FANCJ-R251C and -Q255H were expressed similar to FANCJ-WT, we wanted to determine whether the FANCJ-R251C and -Q255H variants were associated with proteins known to interact with endogenous wildtype FANCJ. Co-immunoprecipitation experiments using nuclear extracts from HeLa cells demonstrated that TopBP1, MLH1, and BRCA1 proteins previously shown to interact with FANCJ (17,31,34) were pulled down by a GFP antibody from lysates of cells that expressed GFP-tagged FANCJ (wild-type and mutants) but not GFP alone (Fig. 9D), suggesting that FANCJ-R251C and -Q255H interact with their protein partners in a manner similar to FANCJ-WT.…”

Mismatch repair protein hMSH2–hMSH6 recognizes mismatches and forms sliding clamps within a D-loop recombination intermediate

“…Msh2 localizes to recombination intermediates in vivo (62), and subsequent heteroduplex rejection during single-strand annealing requires the Sgs1 helicase and Msh2-Msh6 (30). Moreover, the human RecQ family helicases BLM and WRN (63,64) and FANCJ helicase (65) physically and functionally interact with the MMR-components. And a single G/T mismatch located ahead of the fork junction increased the efficiency of the hMSH2-hMSH6-dependent unwinding by WRN helicase (63).…”