R01CA314254

One-ended break repair in mammalian cells - Project summary/abstract

DNA repair initiated at sites of replication fork breakage is critical for the prevention of genomic instability in cycling cells.

Defects in the repair of stressed replication forks have been directly implicated in cancer, notably in hereditary breast and ovarian cancer (HBOC).

We have developed innovative tools for quantifying error-free homologous recombination (HR) and error-prone HR at broken mammalian replication forks or at double strand breaks (DSBs) where completion of HR occurs in an error-prone fashion.

Collectively, the types of HR that this proposal addresses could be characterized as ‘one-ended’.

This phrase means either that a DSB arises as a one-ended break (e.g., at a broken replication fork), or that only one end of a 2-ended DSB has homology with the donor.

In each case, the mechanisms that enable termination of HR are necessarily error-prone, but are poorly understood.

Our previous studies suggest that such breaks may carry a high risk of misrepair with formation of translocations or other chromosome rearrangements, thereby fostering genomic instability and promoting tumorigenesis.

The major goals of this proposal are to define whether mechanisms exist to limit genomic instability at sites of replication fork breakage and one-ended HR in mammalian cells.

We have developed an array of cutting-edge tools to support this study, including unique, sophisticated HR reporters that can distinguish between error-free ‘short tract’ HR and error-prone ‘long tract’ HR—a replicative response likely analogous to break-induced replication.

We have developed a new HR reporter in which HR is triggered by a DNA-protein crosslink (DPC)-induced DNA nick highly similar to the DPC nick induced by the topoisomerase I inhibitor and cancer therapeutic, camptothecin.

This tool can be used to induce fork breakage in a strand-specific manner, enabling us to target breaks to the leading or lagging strand of the approaching replication fork.

We have also developed new tools to analyze the fate of the HR reaction if HR termination occurs without the presence of a homologous second end.

In support of these studies, we will use cutting-edge single molecule ‘long read’ nanopore sequencing to identify and analyze gene conversion tracts and complex breakpoints that arise as a result of HR at one-ended breaks.

Success in this project will reveal the mechanisms that regulate HR repair of broken forks and the mechanisms governing repair pathway “choice” at sites of aberrant HR termination.

This work may also identify new molecular targets for cancer therapy.

Beth Israel Deaconess Medical Center

<P>THE GOALS ARE:</P><UL><LI>TO FOSTER FUNDAMENTAL CREATIVE DISCOVERIES, INNOVATIVE RESEARCH STRATEGIES, AND THEIR APPLICATIONS AS A BASIS FOR ULTIMATELY PROTECTING AND IMPROVING HEALTH;</LI><LI>TO DEVELOP, MAINTAIN, AND RENEW SCIENTIFIC HUMAN AND PHYSICAL RESOURCES THAT WILL ENSURE THE NATION'S CAPABILITY TO PREVENT DISEASE;</LI><LI>TO EXPAND THE KNOWLEDGE BASE IN MEDICAL AND ASSOCIATED SCIENCES IN ORDER TO ENHANCE THE NATION'S ECONOMIC WELL-BEING AND ENSURE A CONTINUED HIGH RETURN ON THE PUBLIC INVESTMENT IN RESEARCH; AND</LI><LI>TO EXEMPLIFY AND PROMOTE THE HIGHEST LEVEL OF SCIENTIFIC INTEGRITY, PUBLIC ACCOUNTABILITY, AND SOCIAL RESPONSIBILITY IN THE CONDUCT OF SCIENCE.</LI></UL><P>IN REALIZING THESE GOALS, THE NIH PROVIDES LEADERSHIP AND DIRECTION TO PROGRAMS DESIGNED TO IMPROVE THE HEALTH OF THE NATION BY CONDUCTING AND SUPPORTING RESEARCH:</P><UL><LI>IN THE CAUSES, DIAGNOSIS, PREVENTION, AND CURE OF HUMAN DISEASES;</LI><LI>IN THE PROCESSES OF HUMAN GROWTH AND DEVELOPMENT;</LI><LI>IN THE BIOLOGICAL EFFECTS OF ENVIRONMENTAL CONTAMINANTS;</LI><LI>IN THE UNDERSTANDING OF MENTAL, ADDICTIVE AND PHYSICAL DISORDERS; AND</LI><LI>IN DIRECTING PROGRAMS FOR THE COLLECTION, DISSEMINATION, AND EXCHANGE OF INFORMATION IN MEDICINE AND HEALTH, INCLUDING THE DEVELOPMENT AND SUPPORT OF MEDICAL LIBRARIES AND THE TRAINING OF MEDICAL LIBRARIANS AND OTHER HEALTH INFORMATION SPECIALISTS.</LI></UL>

93.393 - Cancer Cause and Prevention Research

National Cancer Institute (NCI) [HHS - NIH]

Boston, Massachusetts 022155400 United States

Single Zip Code

NIH Research Project Grant (Parent R01 Clinical Trial Not Allowed) (PA-25-301)