Kaposi's Sarcoma-Associated Herpesvirus Viral Interleukin-6 Signaling Upregulates Integrin ß3 Levels and Is Dependent on STAT3.

Kaposi's sarcoma-associated herpesvirus (KSHV) is the causative agent of two B-cell lymphoproliferative diseases and Kaposi's sarcoma, an endothelial-cell-driven cancer. KSHV viral interleukin-6 (vIL-6) is a viral homolog of human IL-6 (hIL-6) that is expressed in KSHV-associated malignancies. Previous studies have shown that the expression of the integrin ß3 (ITGB3) subunit is induced upon KSHV infection. Here we report that KSHV vIL-6 is able to induce the expression of ITGB3 and increase surface expression of the αVß3 integrin heterodimer. We demonstrated using small interfering RNA (siRNA) depletion and inhibitor studies that KSHV vIL-6 can increase ITGB3 by inducing STAT3 signaling. Furthermore, we found that secreted vIL-6 is capable of inducing ITGB3 in endothelial cells in a paracrine manner. Importantly, the ability to induce ITGB3 in endothelial cells seems to be specific to vIL-6, as overexpression of hIL-6 alone did not affect levels of this integrin. Our lab and others have previously shown that vIL-6 can induce angiogenesis, and we investigated whether ITGB3 was involved in this process. We found that siRNA depletion of ITGB3 in vIL-6-expressing endothelial cells resulted in a decrease in adhesion to extracellular matrix proteins. Moreover, depletion of ITGB3 hindered the ability of vIL-6 to promote angiogenesis. In conclusion, we found that vIL-6 can singularly induce ITGB3 and that this induction is dependent on vIL-6 activation of the STAT3 signaling pathway.IMPORTANCE Kaposi's sarcoma-associated herpesvirus (KSHV) is the etiological agent of three human malignancies: multicentric Castleman's disease, primary effusion lymphoma, and Kaposi's sarcoma. Kaposi's sarcoma is a highly angiogenic tumor that arises from endothelial cells. It has been previously reported that KSHV infection of endothelial cells leads to an increase of integrin αVß3, a molecule observed to be involved in the angiogenic process of several malignancies. Our data demonstrate that the KSHV protein viral interleukin-6 (vIL-6) can induce integrin ß3 in an intracellular and paracrine manner. Furthermore, we showed that this induction is necessary for vIL-6-mediated cell adhesion and angiogenesis, suggesting a potential role of integrin ß3 in KSHV pathogenesis and development of Kaposi's sarcoma.

Functional Comparison of HBZ and the Related APH-2 Protein Provides Insight into Human T-Cell Leukemia Virus Type 1 Pathogenesis.

UNLABELLED: Human T-cell leukemia virus type 1 (HTLV-1) and type 2 (HTLV-2) are highly related retroviruses that transform T cells in vitro but have distinct pathological outcomes in vivo. HTLV-1 encodes a protein from the antisense strand of its proviral genome, the HTLV-1 basic leucine zipper factor (HBZ), which inhibits Tax-1-mediated viral transcription and promotes cell proliferation, a high proviral load, and persistence in vivo. In adult T-cell leukemia/lymphoma (ATL) cell lines and patient T cells, hbz is often the only viral gene expressed. The antisense strand of the HTLV-2 proviral genome also encodes a protein termed APH-2. Like HBZ, APH-2 is able to inhibit Tax-2-mediated viral transcription and is detectable in most primary lymphocytes from HTLV-2-infected patients. However, unlike HBZ, the loss of APH-2 in vivo results in increased viral replication and proviral loads, suggesting that HBZ and APH-2 modulate the virus and cellular pathways differently. Herein, we examined the effect of APH-2 on several known HBZ-modulated pathways: NF-κB (p65) transactivation, transforming growth factor ß (TGF-ß) signaling, and interferon regulatory factor 1 (IRF-1) transactivation. Like HBZ, APH-2 has the ability to inhibit p65 transactivation. Conversely, HBZ and APH-2 have divergent effects on TGF-ß signaling and IRF-1 transactivation. Quantitative PCR and protein half-life experiments revealed a substantial disparity between HBZ and APH-2 transcript levels and protein stability, respectively. Taken together, our data further elucidate the functional differences between HBZ and APH-2 and how these differences can have profound effects on the survival of infected cells and, ultimately, pathogenesis. IMPORTANCE: Human T-cell leukemia virus type 1 (HTLV-1) and type 2 (HTLV-2) are highly related retroviruses that have distinct pathological outcomes in infected hosts. Functional comparisons of HTLV-1 and HTLV-2 proteins provide a better understanding about how HTLV-1 infection is associated with disease and HTLV-2 infection is not. The HTLV genome antisense-strand genes hbz and aph-2 are often the only viral genes expressed in HTLV-infected T cells. Previously, our group found that HTLV-1 HBZ and HTLV-2 APH-2 had distinct effects in vivo and hypothesized that the differences in the interactions of HBZ and APH-2 with important cell signaling pathways dictate whether cells undergo proliferation, apoptosis, or senescence. Ultimately, these functional differences may affect how HTLV-1 causes disease but HTLV-2 generally does not. In the current study, we compared the effects of HBZ and APH-2 on several HTLV-relevant cellular pathways, including the TGF-ß signaling, NF-κB activation, and IRF-1 transactivation pathways.

Human T-cell leukemia virus type 2 antisense viral protein 2 is dispensable for in vitro immortalization but functions to repress early virus replication in vivo.

Human T-cell leukemia virus type 1 (HTLV-1) and HTLV-2 are closely related but pathogenically distinct human retroviruses. The antisense strand of the HTLV-1 genome encodes HTLV-1 basic leucine zipper (b-ZIP) protein (HBZ), a protein that inhibits Tax-mediated viral transcription, enhances T-cell proliferation, and promotes viral persistence. Recently, an HTLV-2 antisense viral protein (APH-2) was identified. Despite its lack of a typical b-ZIP domain, APH-2, like HBZ, interacts with cyclic AMP response element binding protein (CREB) and downregulates Tax-mediated viral transcription. Here, we provide evidence that the APH-2 C-terminal LXXLL motif is important for CREB binding and Tax repression. In order to investigate the functional role of APH-2 in the HTLV-2-mediated immortalization of primary T lymphocytes in vitro and in HTLV-2 infection in vivo, we generated APH-2 mutant viruses. In cell cultures, the immortalization capacities of APH-2 mutant viruses were indistinguishable from that of wild-type HTLV-2 (wtHTLV-2), indicating that, like HBZ, APH-2 is dispensable for viral infection and cellular transformation. In vivo, rabbits inoculated with either wtHTLV-2 or APH-2 mutant viruses established a persistent infection. However, the APH-2 knockout virus displayed an increased replication rate, as measured by an increased viral antibody response and a higher proviral load. In contrast to HTLV-1 HBZ, we show that APH-2 is dispensable for the establishment of an efficient infection and persistence in a rabbit animal model. Therefore, antisense proteins of HTLV-1 and HTLV-2 have evolved different functions in vivo, and further comparative studies will provide fundamental insights into the distinct pathobiologies of these two viruses.

Recent advances in understanding Kaposi's sarcoma-associated herpesvirus.

Kaposi's sarcoma (KS)-associated herpesvirus (KSHV) is an oncogenic human herpesvirus. KSHV is associated with three cancers in the human population: KS, primary effusion lymphoma (PEL), and multicentric Castleman's disease (MCD). KS is the leading cause of cancer in HIV-infected individuals. In this review, we discuss the most recent discoveries behind the mechanisms of KSHV latency maintenance and lytic replication. We also review current therapies for KSHV-associated cancers.

Identification and characterization of HTLV-1 HBZ post-translational modifications.

Human T-cell leukemia virus type-1 (HTLV-1) is estimated to infect 15-25 million people worldwide, with several areas including southern Japan and the Caribbean basin being endemic. The virus is the etiological agent of debilitating and fatal diseases, for which there is currently no long-term cure. In the majority of cases of leukemia caused by HTLV-1, only a single viral gene, hbz, and its cognate protein, HBZ, are expressed and their importance is increasingly being recognized in the development of HTLV-1-associated disease. We hypothesized that HBZ, like other HTLV-1 proteins, has properties and functions regulated by post-translational modifications (PTMs) that affect specific signaling pathways important for disease development. To date, PTM of HBZ has not been described. We used an affinity-tagged protein and mass spectrometry method to identify seven modifications of HBZ for the first time. We examined how these PTMs affected the ability of HBZ to modulate several pathways, as measured using luciferase reporter assays. Herein, we report that none of the identified PTMs affected HBZ stability or its regulation of tested pathways.