The study of homologous and nonhomologous recombination in animal cells is in its infancy. For the near future, the genetic analysis of these processes will involve construction of increasingly sophisticated DNA substrates, which can be introduced into cells to probe in detail their recombinational capabilities. Transfection of modified SV40 genomes into monkey cells offers an especially powerful approach to these studies 1) because the input genomes can be manipulated to distinguish among potential mechanisms, and 2) because the products of recombination are confined within the same plaque, allowing examination of the DNA segments that interacted in the exchange. This latter advantage is unique to the SV40 system. Experiments in this system and others have begun to outline the mechanisms of homologous and nonhomologous recombination in animal cells. In this application we propose to extend our characterization of these processes in monkey cells. Our experiments focus on specific mechanistic steps that are amenable to attack using the SV40 system; in several cases they will provide critical tests of the models we and others have put forward. Our specific objectives are summarized under the more general questions listed below. 1. Are regions of single-stranded DNA involved in the breakage and joining steps of nonhomologous recombination? 2. Are mismatches in heteroduplex intermediates in homologous recombination repaired efficiently and can the enzymatic activities be isolated? 3. Are both potential products or only one preserved in individual homologous exchange? 4. What is the basis for the competition between homologous and nonhomologous recombination and can it be controlled?

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
2R01CA015743-13
Application #
3164248
Study Section
Experimental Virology Study Section (EVR)
Project Start
1979-08-01
Project End
1988-11-30
Budget Start
1986-02-01
Budget End
1986-11-30
Support Year
13
Fiscal Year
1986
Total Cost
Indirect Cost
Name
Baylor College of Medicine
Department
Type
Schools of Medicine
DUNS #
074615394
City
Houston
State
TX
Country
United States
Zip Code
77030
Roth, D B; Chang, X B; Wilson, J H (1989) Comparison of filler DNA at immune, nonimmune, and oncogenic rearrangements suggests multiple mechanisms of formation. Mol Cell Biol 9:3049-57
Zheng, H; Chang, X B; Wilson, J H (1989) Primary cells and established cell lines join DNA ends with the same efficiency relative to homologous recombination. Plasmid 22:99-105
Weiss, U; Wilson, J H (1989) Effects of nicks on repair of single-stranded loops in heteroduplex DNA in mammalian cells. Somat Cell Mol Genet 15:13-8
Weiss, U; Wilson, J H (1988) Heteroduplex-induced mutagenesis in mammalian cells. Nucleic Acids Res 16:2313-22
Chang, X B; Wilson, J H (1987) Modification of DNA ends can decrease end joining relative to homologous recombination in mammalian cells. Proc Natl Acad Sci U S A 84:4959-63
Weiss, U; Wilson, J H (1987) Repair of single-stranded loops in heteroduplex DNA transfected into mammalian cells. Proc Natl Acad Sci U S A 84:1619-23
Chang, X B; Wilson, J H (1986) Formation of deletions after initiation of simian virus 40 replication: influence of packaging limit of the capsid. J Virol 58:393-401
Roth, D B; Wilson, J H (1986) Nonhomologous recombination in mammalian cells: role for short sequence homologies in the joining reaction. Mol Cell Biol 6:4295-304
Roth, D B; Wilson, J H (1985) Relative rates of homologous and nonhomologous recombination in transfected DNA. Proc Natl Acad Sci U S A 82:3355-9
Wake, C T; Vernaleone, F; Wilson, J H (1985) Topological requirements for homologous recombination among DNA molecules transfected into mammalian cells. Mol Cell Biol 5:2080-9

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