Characterization of MRE11 Mutants Provides Insight into Human Genetic Disease, Cancer, and Clinical Intervention.

DNA double strand breaks (DSBs) pose a serious threat to cellular and organism well-being. In response to DSBs, MRE11/RAD50/NBS1 (MRN) initiates DNA repair and facilitates signaling via ataxia-telangiectasia mutated (ATM) and ataxia-telangiectasia- and rad3-related (ATR) kinases. Though alteration of any one of several MRE11 protein interaction partners contributes to human genetic disease and/or carcinogenesis, the role MRE11A mutation plays in these processes remains unclear. I have set out to clarify this role with potential implications in cancer prevention, prognostication, and treatment. I studied four human disease-associated MRE11 mutants. Mutants had altered phosphodiesterase domain (MRE11 W243R (ATLD17) and MRE11 Del340-366 (ASM)), glycine-arginine-rich (GAR) motif (MRE11 R572Q (GRM)), or cyclin-dependent kinase (CDK) 2-interacting motif (MRE11 R633X (ATLD1)). Murine versions of the mutants were expressed to physiologic MRE11 levels in murine embryonic fibroblasts, and conditional deletion abolished wild-type MRE11 expression. In this context, I assessed the mutant proteins’ abilities to facilitate DSB response (DSBR) signaling. Each cancer-associated mutant had distinct effects on ionizing radiation (IR)-induced DSBR. ATLD17, ASM, and GRM were defective in facilitating ATM, ATM and ATR, and ATR activity, respectively. Only the impact of ASM obviously disabled the G2/M checkpoint. No defect in ATLD1-facilitated kinase activity was detected. Whereas ATLD17- and ASM-associated deficits appeared to be attributable to varying disruptions in the MRN complex, no such disruptions were evident with GRM or ATLD1. MRE11 functional roles clarified include those of the phosphodiesterase domain and homodimerization motif in ATM activation and those of the MRE11 GAR and CDK2-interacting motifs in DSBR signaling. Physiologic MRE11 levels were found to be crucial for optimal DSBR signaling. Because our findings concern the influences of MRE11 protein levels, protein folding, protein-protein interactions, and post-translational modifications on fundamental cellular processes, these findings may broadly inform understanding of MRE11 and protein complexes containing MRE11 in normal and disease states.PhDMolecular and Cellular PathologyUniversity of Michigan, Horace H. Rackham School of Graduate Studieshttp://deepblue.lib.umich.edu/bitstream/2027.42/111392/1/regalja_1.pd