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Studies of retroviral vectors for in utero gene transfer and investigation of calcium-mediated gene regulation by Human T-lymphotropic virus type-1

Nair, Amrithraj Muraleedharan
http://rave.ohiolink.edu/etdc/view?acc_num=osu1088785797

Abstract Details

2004, Doctor of Philosophy, Ohio State University, Veterinary Biosciences.
Retrovirus-derived vectors provide efficient means of gene-transfer and are potential tools for therapeutic intervention against congenital or inherited disorders by in-utero gene therapy and for gene-transfer into dividing cells both in vitro and in vivo. To improve biodistribution of retroviral gene-transfer vectors and to obtain efficient gene-transfer towards hematopoietic stem cells via pre-natal administration, we performed in-utero administration of retroviral vectors pseudotyped with different envelope glycoproteins. Relatively high gene-transfer was obtained with significant differences in biodistribution and gene-transfer efficiency using various pseudotypes. There was significant reduction in the percentage of colonies containing transgene, 3 months postnatally. Next, we used HIV-derived lentiviral vectors to investigate the role of human T-lymphotropic virus type-1 (HTLV-1) accessory protein p12(I) in viral pathogenesis. HTLV-1, a deltaretrovirus cause adult T-cell leukemia/lymphoma, and other lymphocyte-mediated disorders. HTLV-1 provirus encodes various regulatory and accessory genes, including ORF-I encoded p12(I). We have demonstrated that this endoplasmic reticulum-localizing protein, increases intracellular calcium, activates NFAT-mediated transcription and is critical for infectivity in vivo and in vitro. Herein, using microarrays, we tested the role of p12(I) in regulating cellular gene expression in T-lymphocytes and found that p12(I) altered the expression of several calcium-regulated genes and genes associated with T-cell signaling, cell-proliferation and apoptosis. Furthermore, p12(I) upregulated the levels of p300 to transcriptionally significant levels, in T-lymphocytes. Next, we further characterized the mechanism of p300 upregulation by p12(I) and demonstrate that it is calcium-dependent, but calcineurin-independent. Sustained low-magnitude calcium release results in increased p300 RNA, protein and p300-mediated transcriptional activity in T-lymphocytes. Calcium-responsive p300-mediated transcription was inhibited by the p300-binding protein, adenovirus E1A, but not by p300-binding deficient E1A mutant. Additionally, ER localization of p12(I) is required for enhancing p300 levels. Collectively, our data indicate that retroviral vectors can be improved to provide effective tools for gene delivery. Using this approach, we found that HTLV-1 p12(I) modulates cellular gene expression to facilitate T-lymphocyte activation and enhances the expression of a rate-limiting co-adaptor p300 in a calcium-dependent manner. These findings provide valuable insights into the mechanisms of HTLV-1 mediated T-lymphocyte proliferation to enhance viral infectivity and clonal expansion of its genome during cell division.
Michael Lairmore (Advisor)
296 p.
Biology, Molecular
Retrovirus; In utero; Gene therapy; Pseudotyping; HTLV-1; Accessory protein; p12(I); Calcium; Gene array; Modulate cellular gene expression; p300; Viral pathogenesis

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Citations

  • Nair, A. M. (2004). Studies of retroviral vectors for in utero gene transfer and investigation of calcium-mediated gene regulation by Human T-lymphotropic virus type-1 [Doctoral dissertation, Ohio State University]. OhioLINK Electronic Theses and Dissertations Center. http://rave.ohiolink.edu/etdc/view?acc_num=osu1088785797

    APA Style (7th edition)

  • Nair, Amrithraj. Studies of retroviral vectors for in utero gene transfer and investigation of calcium-mediated gene regulation by Human T-lymphotropic virus type-1. 2004. Ohio State University, Doctoral dissertation. OhioLINK Electronic Theses and Dissertations Center, http://rave.ohiolink.edu/etdc/view?acc_num=osu1088785797.

    MLA Style (8th edition)

  • Nair, Amrithraj. "Studies of retroviral vectors for in utero gene transfer and investigation of calcium-mediated gene regulation by Human T-lymphotropic virus type-1." Doctoral dissertation, Ohio State University, 2004. http://rave.ohiolink.edu/etdc/view?acc_num=osu1088785797

    Chicago Manual of Style (17th edition)

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This open access ETD is published by The Ohio State University and OhioLINK.