Cell-to-cell transmission of retroviruses, such as human T lymphotropic virus type 1 (HTLV-1), is well documented, but the roles of viral regulatory or other nonstructural proteins in the modulation of T cell adhesion are incompletely understood. In this study we tested the role of the HTLV-1 accessory protein, p12I, on LFA-1-mediated cell adhesion. p12I is critical for early HTLV-1 infection by causing the release of calcium from the endoplasmic reticulum to activate NFAT-mediated transcription. We tested the role of this novel viral protein in mediating LFA-1-dependent cell adhesion. Our data indicated that T cells expressing a mutant HTLV-1 provirus that does not produce p12I mRNA (ACH.p12I) exhibited reduced LFA-1-mediated adhesion compared with wild-type HTLV-1-expressing cells (ACH). Furthermore, the expression of p12I in Jurkat T cells using lentiviral vectors enhanced LFA-1-mediated cell adhesion, which was inhibited by the calcium chelator BAPTA-AM, the calcium channel blocker SK&F 96365, and calpeptin, an inhibitor of the calcium-dependent protease calpain. Similar to the intracellular calcium mobilizer, thapsigargin, the expression of p12I in Jurkat T cells induced cell surface clustering of LFA-1 without changing the level of integrin expression. Our data are the first to indicate that HTLV-1 p12I, in addition to enhancing T cell activation, promotes cell-to-cell spread by inducing LFA-1 clustering on T cells via calcium-dependent signaling.

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Human T-lymphotropic virus type-1 (HTLV-1) is a deltaretrovirus that causes adult T cell leukemia/lymphoma, and is implicated in a variety of lymphocyte-mediated inflammatory disorders. HTLV-1 provirus has regulatory and accessory genes in four pX open reading frames. HTLV-1 pX ORF-II encodes two proteins, p13II and p30II, which are incompletely defined in virus replication or pathogenesis. We have demonstrated that pX ORF-II mutations block virus replication in vivo and that ORF-II encoded p30II, a nuclear-localizing protein that binds with CREB-binding protein (CBP)/p300, represses CREB and Tax responsive element (TRE)-mediated transcription. Herein, we have identified p30II motifs important for p300 binding and in regulating TRE-mediated transcription in the absence and presence of HTLV-1 provirus. Within amino acids 100-179 of p30II, a region important for repression of LTR-mediated transcription, we identified a single lysine residue at amino acid 106 (K3) that significantly modulates the ability of p30II to repress TRE-mediated transcription. Exogenous p300, in a dose-responsive manner, reverses p30II-dependent repression of TRE-mediated transcription, in the absence or presence of the provirus, In contrast to wild type p300, p300 HAT mutants (defective in histone acetyltransferase activity) only partially rescued p30(II)-mediated LTR repression. Deacetylation by histone deacetylase-1 (HDAC-1) enhanced p30II-mediated LTR repression, while inhibition of deacetylation by trichostatin A decreases p30(II)-mediated LTR repression. Collectively, our data indicate that HTLV-1 p30II modulates viral gene expression in a cooperative manner with p300-mediated acetylation.

II 's influence on lymphocyte survival. Our data are the first to demonstrate that p13 II alters Ras-mediated apoptosis in T lymphocytes, and they reveal a potential mechanism by which HTLV-1 alters lymphocyte proliferation.

Human T-lymphotropic virus type 1 (HTLV-1) is the etiologic agent of adult T cell leukemia/ lymphoma (ATLL), an aggressive CD4 + T lymphocyte malignancy. Activation of T lymphocytes is required for effective retroviral integration into the host cell genome and subsequent viral replication, but the molecular mechanisms involved in HTLV-1-mediated T cell activation remain unclear. HTLV-1 encodes various accessory proteins such as p12 I , which has been demonstrated to be critical for HTLV-1 infectivity in vivo in rabbits and in vitro in quiescent primary human T lymphocytes. This hydrophobic protein localizes in the endoplasmic reticulum, increases intracellular calcium, and activates nuclear factor of activated T cell-mediated transcription. To further elucidate the role of p12 I in regulation of cellular gene expression, we performed gene array analysis on stable p12 Iexpressing Jurkat T cells, using Affymetrix U133A arrays. Our data indicate that p12 I altered the expression of genes associated with a network of interrelated pathways including T cell signaling, cell proliferation, and apoptosis. Expression of several calcium-regulated genes was found to be altered by p12 I , consistent with known properties of the viral protein. Gene array findings were confirmed by semiquantitative RT-PCR in Jurkat T cells and primary CD4 + T lymphocytes. Furthermore, dose-dependent expression of p12 I in Jurkat T cells resulted in significant increases in p300 and p300-dependent transcription. This is the first report of a viral protein influencing the transcription of p300, a rate-limiting coadapter critical in HTLV-1-mediated T cell activation.