The human T-cell leukemia/lymphotropic virus type 1 p12I proteins bind the interleukin-2 receptor beta and gammac chains and affects their expression on the cell surface.

p12I is a small hydrophobic protein encoded by the human T-cell leukemia/lymphotropic virus type 1 (HTLV-1) that interacts with the 16-kDa component of the H+ vacuolar ATPase and cooperates with bovine papillomavirus 1 E5 oncoprotein in cell transformation. Just as an important step in E5 action appears to be its binding to the platelet-derived growth factor receptor, it was found that p12I binds specifically to both the beta and gamma(c) chains of the interleukin-2 receptor (IL-2R). The IL-2R beta and gamma(c) chains associated with p12I are endoglycosidase-H sensitive, suggesting that their interaction occurs in a pre-Golgi compartment. p12I stabilizes the immature forms of the IL-2R beta and gamma(c) chains and decreases their cell surface expression. The interactions of p12I with IL-2R beta and gamma(c) may have important implications in the immunosuppressive effect of HTLV-1 in vivo as well as in the ligand-independent HTLV-1-mediated T-cell proliferation.     

Identification, using synthetic peptides, of the minimum amino acid sequence from the retroviral transmembrane protein p15E required for inhibition of lymphoproliferation and its similarity to gp21 of human T-lymphotropic virus types I and II.

Synthetic peptides containing portions of a highly conserved region of retroviral transmembrane proteins of human and animal retroviruses were tested for their ability to inhibit lymphoproliferation to determine the minimum amino acid sequence required. The previously reported immunosuppression mediated by the peptide CKS-17 was confirmed and further localized to a sequence of eight residues essentially identical to the sequence present in the transmembrane protein gp21 of human T-lymphotropic virus types I and II (HTLV-I and -II). To substantiate the physiological relevance of the inhibition of lymphoproliferation observed with the synthetic peptides and to relate this activity to the intact protein, we purified the Rauscher murine leukemia virus transmembrane protein p15E by immunoaffinity chromatography and report that this purified component presented in the form of protein micelles inhibited the interleukin-2-dependent proliferation of the murine T-cell line CTLL-2 in a dose-dependent manner, with a half-maximal inhibitory dose (ID50) of approximately 16 nM. In comparison, the ID50 concentration of a recombinant form of p15E required to inhibit lymphoproliferation was approximately 2.2 microM. The results reported here support the hypothesis that the transmembrane protein gp21 of HTLV-I and -II participates in the mechanism of immunosuppression previously reported for the transmembrane proteins of feline leukemia virus and other animal retroviruses. Thus, the transmembrane protein of HTLV-I, the etiological agent of adult T-cell leukemia-lymphoma, may be partially responsible for the immunocompromised clinical course of this disease that results in fatal opportunistic infections in a majority of cases.     

Evidence for cooperative transforming activity of the human pituitary tumor transforming gene and human T-cell leukemia virus type 1 Tax

Aneuploidy is frequent in cancers. Recently it was found that pituitary tumor transforming gene (PTTG; also called Pds1p or securin) is overexpressed in many different tumors. Human T-cell leukemia virus type 1 (HTLV-1) is a retrovirus that primarily infects CD4+ T lymphocytes and causes adult T-cell leukemia. Here, we report that overexpression of human PTTG cooperated with the HTLV-I Tax oncoprotein in cellular transformation. Coexpression of Tax and PTTG enhanced chromosomal instability and neoplastic changes to levels greater than overexpression of either factor singularly. Cells that overexpressed both PTTG and Tax induced tumors more robustly in nude mice than cells that expressed either PTTG alone or Tax alone.     

Bovine papillomavirus E7 oncoprotein inhibits anoikis

The bovine papillomavirus type 1 (BPV-1) E7 oncoprotein is required for the full transformation activity of the virus. Although BPV-1 E7 by itself is not sufficient to induce cellular transformation, it enhances the abilities of the other BPV-1 oncogenes to induce anchorage independence. We have been exploring the mechanisms by which E7 might affect the transformation efficiency of other viral oncoproteins and in particular whether it might protect cells from apoptosis. We report here that BPV-1 E6 and E7 can each independently inhibit anoikis, a type of apoptosis that is induced upon cell detachment. Using site-directed mutagenesis, we determined regions of the E7 protein that were essential for its antiapoptotic activity. The ability of E7 to inhibit anoikis did partially correlate with an ability to enhance anchorage independence of BPV-1 E6-transformed cells. In addition, the antiapoptotic activity of E7 also only partially correlated with its ability to bind p600, a cellular protein that has previously been reported to play a role in anoikis. We conclude that the contribution of E7 to BPV-induced cellular transformation may involve its ability to inhibit anoikis but that additional functional activities must also be involved.     

Inhibition of T-cell receptor signal transduction and viral expression by the linker for activation of T cells-interacting p12(I) protein of human T-cell leukemia/lymphoma virus type 1

The p12(I) protein of human T-cell leukemia/lymphoma virus type 1 (HTLV-1) is a small oncoprotein that increases calcium release following protein kinase C activation by phorbol myristate acetate, and importantly, this effect is linker for activation of T cells (LAT) independent. Here, we demonstrate that p12(I) inhibits the phosphorylation of LAT, Vav, and phospholipase C-gamma 1 and decreases NFAT (nuclear factor of activated T cells) activation upon engagement of the T-cell receptor (TCR) with anti-CD3 antibody. Furthermore, we demonstrate that p12(I) localizes to membrane lipid rafts and, upon engagement of the TCR, relocalizes to the interface between T cells and antigen-presenting cells, defined as the immunological synapse. A p12(I) knockout molecular clone of HTLV-1 expresses more virus upon antigen stimulation than the isogenic wild type, suggesting that, by decreasing T-cell responsiveness, p12(I) curtails viral expression. Thus, p12(I) has contrasting effects on TCR signaling: it down-regulates TCR in a LAT-dependent manner on one hand, and on the other, it increases calcium release in a LAT-independent manner. The negative regulation of T-cell activation by p12(I) may have evolved to minimize immune recognition of infected CD4(+) T cells, to impair the function of infected cytotoxic CD8(+) T cells, and to favor viral persistence in the infected host.     

The human papillomavirus type 16 E5 oncoprotein translocates calpactin I to the perinuclear region

The human papillomavirus type 16 (HPV-16) E5 oncoprotein is embedded in membranes of the endoplasmic reticulum and nuclear envelope with its C terminus exposed to the cytoplasm. Among other activities, E5 cooperates with the HPV E6 oncoprotein to induce koilocytosis in human cervical cells and keratinocytes in vitro. The effect of E5 on infected cells may rely on its interactions with various cellular proteins. In this study we identify calpactin I, a heterotetrameric, Ca(2+)- and phospholipid-binding protein complex that regulates membrane fusion, as a new cellular target for E5. Both the annexin A2 and p11 subunits of calpactin I coimmunoprecipitate with E5 in COS cells and in human epithelial cell lines, and an intact E5 C terminus is required for binding. Moreover, E5-expressing cells exhibit a perinuclear redistribution of annexin A2 and p11 and show increased fusion of perinuclear membrane vesicles. The C terminus of E5 is required for both the perinuclear redistribution of calpactin I and increased formation of perinuclear vacuoles. These results support the hypothesis that the E5-induced relocalization of calpactin I to the perinuclear region promotes perinuclear membrane fusion, which may underlie the development of koilocytotic vacuoles.     

The human papillomavirus type 6 and 16 E5 proteins are membrane-associated proteins which associate with the 16-kilodalton pore-forming protein.

The human papillomavirus (HPV) E5 proteins are predicted from DNA sequence analysis to be small hydrophobic molecules, and the HPV type 6 (HPV-6) and HPV-11 E5 proteins share several structural similarities with the bovine papillomavirus type 1 (BPV-1) E5 protein. Also similar to the BPV-1 E5 protein, the HPV-6 and HPV-16 E5 proteins exhibit transforming activity when assayed on NIH 3T3 and C127 cells. In this study, we expressed epitope-tagged E5 proteins from both the "low-risk" HPV-6 and the "high-risk" HPV-16 in order to permit their immunologic identification and biochemical characterization. While the HPV-6 and HPV-16 E5 proteins fail to form disulfide-linked dimers and oligomers, they did resemble the BPV-1 E5 protein in their intracellular localization to the Golgi apparatus, endoplasmic reticulum, and nuclear membranes. In addition, the HPV E5 proteins also bound to the 16-kDa pore-forming protein component of the vacuolar ATPase, a known characteristic of the BPV-1 E5 protein. These studies reveal a common intramembrane localization and potential cellular protein target for both the BPV and HPV E5 proteins.     

Expression of the gag gene of human T-cell leukemia virus type I in Escherichia coli and its diagnostic use

An expression plasmid, pHY202, was constructed which directs the synthesis of a fusion protein encoded by the gag sequence of human T-cell leukemia virus type I (HTLV-I) inserted into the lacZ' gene. Escherichia coli cells harboring pHY202 produced the 43-kDal LacZ'-Gag fusion protein with a yield of approx. 0.3% of total soluble proteins. The fusion protein is specifically recognized by monoclonal antibodies against the Gag proteins p19 and p24, and could be applicable for the diagnosis of HTLV-I infection, because almost all sera from HTLV-I carriers gave a positive response in the enzyme-linked immunosorbent assay (ELISA) employing the LacZ'-Gag hybrid protein purified by immunoaffinity column chromatography.