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Yeast. retrotransposons. reverse transcriptase. fidelity. DNA repair. chromosome stabilityMy laboratory focuses on the study of mechanisms and consequences of retrotransposon reverse transcription. Retrotransposons are endogenous genetic elements probably present in the genomes of all eukaryotic organisms. Their locations along chromosomes are not fixed and so they have been dubbed "jumping genes" or "mobile DNA". Although their normal cellular functions and origins are unknown. they can cause mutations by jumping into or nearby other genes. They can cause large segments of chromosomes to rearrange. thereby altering genome organization. They can capture and carry cellular genes to new locations. Although they are sometimes viewed as genetic parasites. their ubiquity suggests that they serve as agents of evolutionary change and speciation. Their potential role in the genetics of cancer. aging. and heritable diseases are only beginning to be examined. Retrotransposons are also closely related to HIV and other retroviruses. In particular. both types of elements propagate themselves via the enzyme reverse transcriptase (RT) and then randomly insert into new chromosomal locations. We carry out most of our experiments using the yeast retrotransposon Ty1. Studies of many fundamental genetic processes are more easily accomplished and experimental approaches are better developed in yeast than in more complex organisms such as mammals. The major projects going on in my lab are: 1) To understand the link between retrotransposons and DNA double-strand break repair. We have found that DNA synthesized by retrotransposon reverse transcriptases (termed cDNA) can be targeted to a potentially lethal chromosomal break sites. Defects in various DNA repair pathways are associated with genetic diseases. chromosomal instability. and predispositions to cancer. Our findings link the two distinct fields of retrotransposition and DNA repair. Yeast provides a facile model system to study processes which in mammalian cells are associated with cancer. aging. and congenital abnormalities. 2) To study retrotransposon replication fidelity. Reverse transcriptase is a DNA polymerase that copies genetic information in order to replicate. In the case of HIV reverse transcriptase. the process of replication is error prone. i.e. mutations are created during the copying process. We showed that error prone replication occurs during Ty1 replication. and is therefore not limited to infectious viruses. We are examining the consequences of these types of mutations on the ability of retrotransposons to function and interact with the host cell. 3)To determine the functions of reverse transcriptase by mutational analysis of the Ty1 RT. We have created a unique active site Ty1 RT mutant which can still synthesize DNA but is blocked for a subsequent replication step. We are developing genetic and biochemical methods to study the consequences of this and other mutations within Ty1. 4) To examine the determinants of chromosomal rearrangements in yeast. We have developed an assay to identify chromosomal rearrangements caused by nonhomologous recombination. e.g. translocations. inversions. insertions and deletions. We are using this assay to explore the genetic determinants of chromosome stability. Selected PublicationsGabriel A, Dapprich J, Kunkel M, Gresham D, Pratt SC, Dunham MJ. (2006) Global mapping of transposon location. PLoS Genet. 2(12):e212. Pandey. M.. Patel. S. and Gabriel. A.. (2004). Insights into the role of an active site aspartate in Ty1 reverse transcriptase polymerization. Journal of Biological Chemistry 279:47840-47848. Kim. D.Y.. Kim. T.Y.. Walsh. T. Kobayashi. Y.. Matise. T.. Buyske. S.. and Gabriel. A.. (2004). Widespread RNA editing of embedded Alu elements in the human transcriptome. Genome Research 14:1719-1725. Yu. X. and Gabriel. A.. (2004). Reciprocal translocations in Saccharomyces cerevisiae formed by nonhomologous end joining. Genetics 166:741-751. Wilhelm. F-X.. Wilhelm. M. and Gabriel. A.. (2003). Extension and cleavage of the PPT plus-strand primer by Ty1 reverse transcriptase. Journal of Biological Chemistry 278: 47678-47684. Yu. X. And Gabriel. A.. (2003). Ku-dependent and ku-independent end-joining pathways lead to chromosomal rearrangements during double strand break repair in Saccharomyces cerevisiae. Genetics 163:843-856. Wilhelm. M.. Uzun. O.. Mules. E.H.. Gabriel. A. and Wilhelm. F-X. (2001). Polypurine tract formation by Ty1 RNase H. Journal of Biological Chemistry 276:47695-47701. Uzun. O. and Gabriel. A. (2001). A Ty1 reverse transcriptase active-site aspartate mutation blocks transposition but not polymerization. Journal of Virology 75:337-6347. Yu. X. and Gabriel. A. (1999). Patching broken chromosomes with extranuclear cellular DNA. Molecular Cell 4:873-881. Gabriel. A. and Mules. E.H. (1999). Fidelity of retrotransposon replication. Annals ofthe New York Academy of Science. 870:108-118. Mules. E.H. Uzun. O.. and A. Gabriel. (1998).In vivo Ty1 reverse transcription can generate replication intermediates with untidy ends. Journal of Virology. 72:6490-6503. Mules. E.H. Uzun. O.. and Gabriel . A.(1998) Replication errors during in vivo Ty1 transposition are linked to heterogeneous RNAse H cleavage sites. Molecular andCellular Biology. 18:1094-1104. Kim. J.M.. Vanguri. S.. Boeke. J.D.. Gabriel. A.. and Voytas. D.F. (1998). Transposable elements and genome organization: a comprehensive survey of retrotransposons revealed by the complete Saccharomyces cerevisiae genome sequence. Genome Research. 8:464-478. Sassaman. D.M.. Dombroski. B.A.. Moran. J.V.. Kimberland. M.L.. Naas. T.P.. DeBerardinis. R.J.. Gabriel. A.. Swergold. G.D.. and Kazazian. H.H. (1997). Many human L1 elements are capable of retrotransposition. Nature Genetics. 16:37-43. Gabriel. A. and Voytas. D. (1997). DNA on the move. Trends in Genetics. 13:258-259. |
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