Book of Abstracts: Albany 2007
June 19-23 2007
Assembled Structure of the Mre11/Rad50/DNA Complex from X-ray Solution Scattering and Crystallography: Implications for Repair and Signaling
The Mre11/Rad50 complex (Mre11/Rad50/Nbs1 or MRN in higher eukaryotes) plays essential roles in the biologically critical processes of (I) telomere maintenance, (II) DNA double strand break repair by homologous recombination repair and non-homologous end joining, and (III) initial DNA damage sensing and signaling (1, 2). Thus, MRN acts as an enzymatic effecter in telomere maintenance and DNA break repair, as a DNA damage sensor, and a transducer of critical damage response signals to the cell-cycle checkpoint apparatus.
We have determined new X-ray crystal structures of apo- and DNA bound P. furiosus Mre11 that reveal how dimeric Mre11 directly tethers DNA ends. The DNA bridging Mre11-Mre11 interface is essential for function, because charge substitution mutations that disrupt the hydrophobic homo-dimeric surface in vivo and in vitro also ablate Mre11 DNA binding activity, and confer radiation and clastogen sensitivity in S. pombe. To further clarify architectural and enzymatic functions of the Mre11/Rad50 core complex and understand mechanisms of chemo-mechanical communication between Mre11 and Rad50, we have analyzed crystal structures of Rad50 bound to a minimal Mre11 fragment, and dissected structural determinants of MR complex quaternary assembly in solution using Small angle X-ray Scattering (SAXS). SAXS and X-ray structures suggest the Mre112Rad502 core DNA binding head assembles as two juxtaposed half-rings, with dimeric Mre11 wrapped by the two Rad50 coiled-coils, and the Rad50 ATPase domains capping the opposite end of the ring.
These data provide detailed composite structural models of Mre11/Rad50/DNA and suggest novel roles for RAD50 ATP driven conformational controls (4) in DNA break repair and signalling initiation through the ataxia-telangiectasia mutated (ATM) kinase (2). Our observations unify the roles of Mre11 in mediating nuclease-dependent and -independent activities of the MRN complex (5, 6). Higher order MRN architectures and complexes appear appropriate for dictating MRN enzymatic and DNA bridging functions in DNA double strand break repair by homologous recombination repair, non-homologous end joining, and micro-homology mediated DNA repair. MRN linked conformational changes upon ATP and DNA binding provide keys to sensing and signalling upstream of p53.
References and Footnotes
R. Scott Williams1
1Department of Molecular Biology and the Skaggs Institute for Chemical Biology