We have discovered a new family of retroelements, designated diversity-generating retroelements (DGRs) that function to diversify protein-encoding DNA sequences. The prototype DGR was identified in a bacteriophage genome on the basis of its ability to generate variability in a gene that specifies tropism for receptor molecules on Bordetella species. Bordetella cause respiratory infections in humans and other mammals. Tropism switching is a template-dependent, reverse transcriptase-mediated process that introduces nucleotide substitutions at defined locations within a target gene. This cassette-based mechanism is theoretically capable of generating trillions of different amino acid sequences in a single polypeptide, providing a vast repertoire of potential ligand-receptor interactions. Using the Bordetella phage DGR as a signature, we have identified homologous elements in numerous bacterial genomes. Of particular note are DGRs that are predicted to diversify proteins on the surface of Bacteroides thetaiotaomicron, one of the most abundant members of the gastrointestinal flora, and Treponema denticola, an oral bacterium associated with periodontal disease. Prokaryotic DGRs represent an entirely novel mechanism for generating protein diversity. In addition to their fundamental importance as a newly discovered family of genetic elements and their potential roles in human health and disease, DGRs are of interest as a result of their potential applications.
Our specific aims are to: 1. Conduct a mechanistic analysis of the prototype diversity-generating retroelement present in Bordetella bacteriophage. 2. Probe the receptor repertoire and the structural basis of ligand recognition by a DGR-encoded receptor protein. 3. Investigate DGR function in the human gastrointestinal commensal Bacteroides thetaiotaomicron.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
Research Project (R01)
Project #
5R01AI071204-05
Application #
7902305
Study Section
Bacterial Pathogenesis Study Section (BACP)
Program Officer
Khambaty, Farukh M
Project Start
2006-08-01
Project End
2012-07-31
Budget Start
2010-08-01
Budget End
2012-07-31
Support Year
5
Fiscal Year
2010
Total Cost
$350,368
Indirect Cost
Name
University of California Los Angeles
Department
Microbiology/Immun/Virology
Type
Schools of Medicine
DUNS #
092530369
City
Los Angeles
State
CA
Country
United States
Zip Code
90095
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Dai, Wei; Hodes, Asher; Hui, Wong H et al. (2010) Three-dimensional structure of tropism-switching Bordetella bacteriophage. Proc Natl Acad Sci U S A 107:4347-52
Shrivastava, Ruchi; Miller, Jeff F (2009) Virulence factor secretion and translocation by Bordetella species. Curr Opin Microbiol 12:88-93
Guo, Huatao; Tse, Longping V; Barbalat, Roman et al. (2008) Diversity-generating retroelement homing regenerates target sequences for repeated rounds of codon rewriting and protein diversification. Mol Cell 31:813-23
Miller, Jason L; Le Coq, Johanne; Hodes, Asher et al. (2008) Selective ligand recognition by a diversity-generating retroelement variable protein. PLoS Biol 6:e131
Medhekar, Bob; Miller, Jeff F (2007) Diversity-generating retroelements. Curr Opin Microbiol 10:388-95