‘Recombomice’ – A new tool for studying homologous recombination in vivo

Bevin Engelward
Division of Biological Engineering, MIT, Cambridge, MA
Email: bevin@MIT.EDU
Web site: http://web.mit.edu/be/people/engelward.htm

Why this Project?

This project provides a tool (recombomice) for studying homologous recombination. Homology directed repair is critical for genomic stability. However, radiation-induced misalignments and exchanges can lead to deleterious deletions, insertions and loss of heterozygosity events that contribute to cancer. The project will study the role of genetic recombination on radiation-induced cancer.

Project Goals

1. To define the dose range at which homologous recombination occurs in vivo.
2. To reveal the molecular basis for radiation-induced suppression of homologous recombination in vivo

Experimental Approach

1. ‘Recombomice’ have been developed in which recombinant cells fluoresce yellow. For the first time, direct in vivo detection of recombinant cells in somatic tissue is possible. The Fluorescent Yellow Direct Repeat (FYDR) mice carry an engineered direct repeat recombination substrate that yields expression of enhanced yellow fluorescent protein (EYFP) following homologous recombination. These mice provide a remarkably sensitive assay for environmental, genetic and epigenetic factors that affect recombination.
   
2. Fluorescence microscopy and histology will be used to detect recombinant cells within intact pancreatic tissues to study the relative susceptibility of various pancreatic cell types to genetic and environmental factors such as radiation in modulating homologous recombination.

Outcomes

1. Levels of several key DNA repair proteins known to facilitate clearance of recombinogenic DNA lesions are up-regulated in the chronically irradiated animals. These include Polymerase β [Pol β ] and Ku70, which are both required for non-homologous end-joining.
   
2. Chronic irradiation also suppresses the frequency of homologous recombination events to levels below those of unexposed animals. These results are consistent with previous studies showing that ionizing radiation induces a potentially protective adaptive response. The mechanisms involved in these adaptive responses will be studied and their role in radiation-induced cancer defined.