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HIV-1 replication. gene therapy. and retroviral vectorsResearch in our laboratory centers upon studying HIV-1 replication and upon the design and use of retroviral vectors for somatic cell gene therapy involving cells of the lymphoid lineage. Our recent HIV-1 studies have focused upon investigating HIV-1 reverse transcription and recombination. DNA primer strand transfers are essential for HIV-1 replication. and recombination influences the rapid evolution of HIV- 1. The accepted mechanism of retroviral reverse transcription proposes that two primer strand transfers occur during replication. Given the diploid nature of the retroviral virion. the question arises concerning the nature of these primer strand transfers. namely. whether they are intra- or intermolecular. A system was developed to examine the nature of primer transfer events during HIV-1 replication. Both intra- and intermolecular primer transfer were observed. and occurred with similar frequencies during minus strand synthesis. Plus-strand primer transfer was primarily intramolecular. A high rate of homologous recombination occurred during minus-strand DNA synthesis. Recombination occurred at a rate of 3 cross-overs per genome per replication cycle. These results suggest that both viral genomic RNAs serve as templates during HIV-1 reverse transcription and that primer strand transfers as well as recombination contribute to the rapid genetic variation of HIV-1. Primary lymphocytes play a key role in immune surveillance and regulation. Moreover. they are long-lived and easy to obtain. Because of these considerations. they are potentially important targets for somatic cell gene therapy. Using retroviral vectors. we have developed efficient gene transfer procedures for introducing genes into primary lymphocytes. The procedures do not perturb the primary lymphocytes. since after transplantation into recipient hosts. they home to the appropriate lymphoid organs. persist for long periods of time. and express the transferred genes at significant levels. Thus. we are in a position to genetically manipulate lymphocytes and are using this technology to develop procedures for treating autoimmune diseases. such as multiple sclerosis. Selected PublicationsMukherjee S, Lee HL, Pacchia AL, Ron Y, Dougherty JP. (2007) A HIV-2-based self-inactivating vector for enhanced gene transduction. J Biotechnol. 127(4):745-57. Mukherjee S, Lee HL, Ron Y, Dougherty JP. (2006) Proviral progeny of heterodimeric virions reveal a high crossover rate for human immunodeficiency virus type 2. J Virol. 80(24):12402-7. Zhuang J, Mukherjee S, Ron Y, Dougherty JP. (2006) High rate of genetic recombination in murine leukemia virus: implications for influencing proviral ploidy. J Virol. 80(13):6706-11. Micheva-Viteva S. Pacchia AL. Ron Y. Peltz SW. Dougherty JP. (2005) Human immunodeficiency virus type 1 latency model for high-throughput screening. Antimicrob Agents Chemother. 49(12):5185-8. Duttagupta R. Tian B. Wilusz CJ. Khounh DT. Soteropoulos P. Ouyang M. Dougherty JP. Peltz SW. (2005) Global analysis of Pub1p targets reveals a coordinate control of gene expression through modulation of binding and stability. Mol Cell Biol. 25(13):5499-513. Le Roy F. Salehzada T. Bisbal C. Dougherty JP. Peltz SW. (2005) A newly discovered function for RNase L in regulating translation termination. Nat Struct Mol Biol. 12(6):505-12. Biswas. P. Jiang. X. Pacchia,AL. Dougherty. JP and Peltz. SW. (2004) The human immunodeficiency virus type 1 ribosomal frameshifting site is an invariant sequence determinant and an important target for antiviral therapy. J Virol. 78(4):2082-7. Chen CC. Rivera A. Dougherty JP. Ron Y. (2004) Feb 26 [Epub ahead of print] Complete protection from relapsing experimental autoimmune encephalomyelitis induced by Syngeneic B cells expressing the autoantigen. Blood. Adelson. M.. Pacchia. A.L. Kaul. M.. Rando. R.F.. Ron. Y. . Peltz . S.W. and Dougherty. J.P. (2003). Towards the development of a virus/cell based assay for the discovery of novel compounds against HIV-1. Antimicrobial Agents and Chemotherapy 47:501-508. Pacchia. A.L.. Mukherjee. S. and Dougherty. J.P. (2003). Choice and use of appropriate lentiviral packaging cell types. Lentivirus Gene Engineering Protocols. Methods in Molecular Biology. Humana Press Inc. 229:9-42. Oertel. M.. Rosencrantz. R.. Chen. Y.-Q.. Thota. P.N.. Sandhu. J.S.. Dabeva. M..D.. Pacchia. A.L.. Adelson. M.E.. Dougherty. J.P. and Shafritz. D.A. (2003) Repopulation of rat liver by fetal hepatoblasts and adult hepatocytes transduced ex vivo with lentiviral vectors. Hepatology 37(5):994-1005. Zhuang. J.. Jetzt. A. Sun. G. . Yu. H.. Klarmann. G.. Ron. Y.. Preston B.D.. and Dougherty. J.P. (2002). Human immunodeficiency virus type-1 recombination: rate. fidelity. and putative hotspots. Journal of Virology 76:11273-11282. O'Neill. P.. Sun. G.-L. . Yu. H.. Ron. Y.. Dougherty. JP and Preston. B. (2002). Mutational analysis of HIV-1 long terminal repeats to explore the relative contributions of reverse transcriptase and RNA polymerase II to viral mutagenesis. Journal of Biological Chemistry 277:38053-38061. Pacchia. A.L.. Adelson. M.. Ron. Y. and Dougherty. J.P. (2001). A safe lentiviral packaging cell line devoid of the viral accessory genes. Virology 282:77-86. Chen. C.-C.. Rivera. A.. Dougherty. J.P. and Ron. Y. (2001). A gene therapy approach for treating T-cell-mediated autoimmune diseases. Blood 97(4):886-894 . Sun. G.-L.. O'Neill. P.. Yu. H.. Preston. B.. Ron. Y. and Dougherty. J.P. (2001). Transduction of cellular sequences by Human Immunodeficiency Virus Type 1. Journal of Virology 75:11902-11906. |
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