BACH1/FANCJ Checkpoint, Recombination, and Chemoresistance

描述(申请人提供)：化疗耐药性是癌症治疗中的一个主要问题，但很大程度上还不清楚癌细胞是如何产生耐药性的。错配修复(MMR)蛋白的缺失使细胞能够抵抗有毒的化疗药物，如DNA甲基化子。然而，鉴于MMR的多种功能，使细胞对DNA甲基化敏感的不同的MMR功能还没有明确的定义。MMR蛋白的功能是修复DNA复制后的碱基错配，抑制不同DNA序列之间的重组，以及在DNA损伤后激活检查点和凋亡反应。功能分离突变体的研究表明，对DNA甲基化的抗性依赖于一个被破坏的检查点。最近，我们发现MutL1复合体的MMR蛋白(MLH1/PMS2)和BACH1/FANCJ(BRCA1相关的C末端螺旋酶/Fanconi贫血互补组J)都是检查点信号所必需的。具体地说，我们发现BACH1直接与MLH1结合，而为MLH1结合而被去除的BACH1的突变版本未能引起链间交联(ICL)诱导的检查点反应。由于ICL激活了S期内检查点，并且MLH1和BACH1都已被证明在S期内检查点发挥作用，因此该检查点可能需要形成BACH1/MutL1复合体。我们将直接测试这种可能性。此外，我们还将确定BACH1是否也参与了DNA甲基化诱导的类似于MutL1的G2/M积聚检查点。与BACH1在DNA甲基化诱导的反应中所扮演的角色一致，我们的实验室已经表明，类似于MutL1缺陷细胞，BACH1缺陷细胞对DNA甲基化具有抵抗力。相反，BRCA1缺陷细胞对DNA甲基化敏感，这表明BACH1在DNA甲基化反应中具有独特的功能。我们建议剖析BACH1/MutL1复合体在检查点和修复功能中的作用。我们将确定是否需要形成完整的复合体来恢复耐药空BACH1或MutL1细胞的化疗敏感性。理想情况下，确定BACH1/MutL1复合体的功能将为恢复癌细胞的化疗敏感性提供洞察力。沿着这些思路，我们将测试是否操纵BACH1/BRCA1复合体的重组功能将独特地使MMR缺陷细胞对化疗敏感。

DESCRIPTION (provided by applicant): Chemoresistance is a major problem in cancer therapy, but it is largely unclear how cancer cells become resistant. Deficiency in the mismatch-repair (MMR) proteins allows cells to resist toxic chemotherapeutic agents such as DNA methylators. However, the distinct MMR function that sensitizes cells to DNA methylation is not clearly defined given the multiple MMR functions. MMR proteins function to repair base mismatches after DNA replication, inhibit recombination between non-identical DNA sequences, as well as activate both checkpoint and apoptotic responses following DNA damage. Separation-of- function mutants, suggest that resistance to DNA methylation is dependent on a disrupted checkpoint. Recently, we established that both MMR proteins of the MutL1 complex (MLH1/PMS2) and BACH1/FANCJ (BRCA1-associated C- terminal helicase/Fanconi Anemia complementation group J) are required for checkpoint signaling. Specifically, we identified that BACH1 binds directly to MLH1 and that a mutant version of BACH1 ablated for MLH1 binding failed to elicit an interstrand crosslink (ICL)-induced checkpoint response. Since ICLs activate the intra S-phase checkpoint, and both MLH1 and BACH1 have been shown independently to function in the intra S-phase checkpoint, this checkpoint likely requires the formation of a BACH1/MutL1 complex. We will test this possibility directly. In addition, we will determine whether BACH1 also participates in the DNA-methylation-induced G2/M accumulation checkpoint similar to MutL1. Consistent with a role for BACH1 in the DNA methylation-induced response, our lab has shown that similar to MutL1 deficient cells, BACH1 deficient cells are resistant to DNA methylation. In contrast, BRCA1 deficient cells are sensitive to DNA methylation, suggesting that BACH1 uniquely functions in the DNA methylati1n response. We propose to dissect the role of a BACH1/MutL1 complex in both checkpoint and repair functions. We will determine whether the formation of an intact complex is required to restore chemosensitivity to resistant null BACH1 or MutL1 cells. Defining the function of the BACH1/MutL1 complex ideally will provide insight towards restoring chemosensitivity to cancer cells. Along these lines, we will test whether manipulation of the recombination function of the BACH1/BRCA1 complex will uniquely sensitizes MMR deficient cells to chemotherapies.