Abstract

The goal of the proposed research is to generate a family of novel, highly sequence-specific DNA binding proteins with the ability to modify the structure or expression of specific genes in human cells. The starting point for the generation of the Gene-Specific-Reagents (GSR's) are existing proteins known as homing endonucleases, that can bind and cleave unique, long (15-40 bp) DNA targets or homing sites with high specificity in vivo. The mechanism by which these homing endonucleases bind to their specific targets suggest that it may be possible to generate GSR's that can bind to any target sequence of human cell DNA with relatively high specificity. Homing endonuclease variants with the ability to modify the structure or functions of specific human genes would provide new opportunities to investigate human gene structure and function. The availability of human GSR's may also allow modification of altered human genes responsible for specific diseases or genes of pathogenic microorganisms including bacteria and viruses, parasites, etc.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
1R01CA088942-01
Application #
6225738
Study Section
Chemical Pathology Study Section (CPA)
Program Officer
Okano, Paul
Project Start
2001-01-16
Project End
2005-12-31
Budget Start
2001-01-16
Budget End
2001-12-31
Support Year
1
Fiscal Year
2001
Total Cost
$202,392
Indirect Cost

  Institution

Name
University of Washington
Department
Pathology
Type
Schools of Medicine
DUNS #
135646524
City
Seattle
State
WA
Country
United States
Zip Code
98195

  Publications

Eklund, Jennifer L; Ulge, Umut Y; Eastberg, Jennifer et al. (2007) Altered target site specificity variants of the I-PpoI His-Cys box homing endonuclease. Nucleic Acids Res 35:5839-50
Sussman, Django; Chadsey, Meg; Fauce, Steve et al. (2004) Isolation and characterization of new homing endonuclease specificities at individual target site positions. J Mol Biol 342:31-41
Chevalier, Brett; Sussman, Django; Otis, Christian et al. (2004) Metal-dependent DNA cleavage mechanism of the I-CreI LAGLIDADG homing endonuclease. Biochemistry 43:14015-26
Chevalier, Brett; Turmel, Monique; Lemieux, Claude et al. (2003) Flexible DNA target site recognition by divergent homing endonuclease isoschizomers I-CreI and I-MsoI. J Mol Biol 329:253-69
Seligman, Lenny M; Chisholm, Karen M; Chevalier, Brett S et al. (2002) Mutations altering the cleavage specificity of a homing endonuclease. Nucleic Acids Res 30:3870-9
Chevalier, Brett S; Kortemme, Tanja; Chadsey, Meggen S et al. (2002) Design, activity, and structure of a highly specific artificial endonuclease. Mol Cell 10:895-905