Ralph Scully

Department of Pathology
Beth Israel Deaconess Medical Center
Division of Hematology-Oncology/Cancer Biology Program
Harvard Institutes of Medicine, Room 923
77 Avenue Louis Pasteur
Boston, MA 02115
Tel: (617) 667-4252
Fax: (617) 667-0980
Email: rscully@bidmc.harvard.edu
4 postdoctoral fellows, 1 graduate student

We are interested in the relationship between recombination and cancer. A large body of evidence links chromosomal double strand breaks (DSB) to cancer predisposition in mammals. Consistent with this, a number of human tumor suppressor genes have been implicated in control of homologous recombination or in other pathways of DSB repair. A major trigger to homologous recombination in cycling cells is the stalling of a DNA polymerase complex on abnormal DNA structure during the S phase of the cell cycle. Circumstantial evidence suggests that the products of the hereditary breast/ovarian cancer predisposition genes, BRCA1 and BRCA2, together with interacting recombination proteins such as Rad51, enforce a potentially error-free form of homologous recombination, in which the neighboring sister chromatid is used as template for repair (“sister chromatid recombination” - SCR). We developed a novel reporter of SCR, triggered by a site-specific chromosome breakage using a rare-cutting restriction endonuclease, and have recently further refined this reporter to facilitate rapid analysis of SCR by flow cytometry. We are using this system to analyze the roles of BRCA1 and BRCA2 in SCR, as well as other important cancer and development-related genes, such as the Bloom’s syndrome gene, the Fanconi Anemia genes and Rad51-related genes. We have recently embarked on a genetic study of SCR in the experimentally tractable chicken B cell lymphoblastoid cell line, DT40.

Double strand breaks provoke a remarkable reaction in chromatin flanking the break, marked by phosphorylation of the histone H2A variant, H2AX, on a C-terminal serine residue (S139) (to form “?-H2AX”). The ?-H2AX tract extends hundreds of kilobases either side of the break. H2AX–/– mice are cancer prone and H2AX–/– cells reveal genomic instability and DSBR defects. We found that H2AX S139 phosphorylation is required for efficient SCR, implying that the chromatin response around a DSB contributes in some way to efficient DSB repair. This surprising discovery suggests that regulation of DSB repair is organized in chromatin, and is not restricted to enzymes acting at the break. We have recently identified the ?-H2AX interacting adaptor protein, MDC1, as a key mediator of H2AX recombination functions. We are currently using biochemical techniques to identify MDC1 interacting proteins that mediate its SCR functions. We are also developing novel cell imaging and chromatin-immunoprecipitation techniques to study dynamic aspects of the cellular response to DSBs.

References:

• Xie A, Hartlerode A, Stucki M, Odate S, Puget N, Kwok A, Nagaraju G, Yan C, Alt FW, Chen J, Jackson SP and Scully R. Distinct Roles of Chromatin-Associated Proteins MDC1 and 53BP1 in Mammalian Double-Strand Break Repair. Mol Cell 2007; 28: 1045-1057.
• Nagaraju G, Odate S, Xie A and Scully R. Differential regulation of short- and long-tract gene conversion between sister chromatids by Rad51C. Mol Cell Biol 2006; 26(21): 8075-8086.
• Xie A, Puget N, Shim I, Odate S, Jarzyna I, Bassing CH, Alt FW and Scully R. Control of sister chromatid recombination by histone H2AX. Mol Cell 2004; 16: 1017-1025.