Collagen IV (COL4A1, COL4A2), a Component of the Viral Biofilm, Is Induced by the HTLV-1 Oncoprotein Tax and Impacts Virus Transmission

Abstract

Human T-cell leukemia virus type 1 (HTLV-1) is the etiologic agent for Adult T-Cell Leukemia/Lymphoma (ATLL) and HTLV-1-Associated Myelopathy/Tropical Spastic Paraparesis (HAM/TSP). HTLV-1 infects CD4+ T-cells via cell-to-cell transmission requiring reorganization of the cytoskeleton and expression of the viral transactivator and oncoprotein Tax. Viruses spread at the virological synapse (VS), a virus-induced specialized cell-cell contact, by polarized budding into synaptic clefts, and by cell surface transfer of viral biofilms (VBs). Since little is known about Tax’s role in formation of the VB, we asked which component of the VB is regulated by Tax and important for HTLV-1 transmission. Collagens are not only structural proteins of the extracellular matrix and basal membrane but also represent an important component of the VB. Here, we report that among the collagens known to be present in VBs, COL4 is specifically upregulated in the presence of HTLV-1 infection. Further, we found that transient expression of Tax is sufficient to induce COL4A1 and COL4A2 transcripts in Jurkat and CCRF-CEM T-cells, while robust induction of COL4 protein requires continuous Tax expression as shown in Tax-transformed T-cell lines. Repression of Tax led to a significant reduction of COL4A1/A2 transcripts and COL4 protein. Mechanistically, luciferase-based promoter studies indicate that Tax activates the COL4A2 and, to a less extent, the COL4A1 promoter. Imaging showing partial co-localization of COL4 with the viral Gag protein in VBs at the VS and transfer of COL4 and Gag to target cells suggests a role of COL4 in VB formation. Strikingly, in chronically infected C91-PL cells, knockout of COL4A2 impaired Gag transfer between infected T-cells and acceptor T-cells, while release of virus-like particles was unaffected. Taken together, we identified COL4 (COL4A1, COL4A2) as a component of the VB and a novel cellular target of Tax with COL4A2 appearing to impact virus transmission. Thus, this study is the first to provide a link between Tax’s activity and VB formation by hijacking COL4 protein functions.

Identification of Differentially Expressed Genes in Ducks in Response to Avian Influenza A Virus Infections

“…Collagens are part of the extracellular matrix and are crucial for structure repair, tensile stress resistance, cell adhesion, migration, cell-cell interactions, and chemotaxis (Millen et al, 2019). AIV infection is known to be accompanied by lesions in tissues where it invades as a result of inflammatory responses and secondary bacterial infection, causing tissue damage.…”

Microbial Biofilms: Human T-cell Leukemia Virus Type 1 First in Line for Viral Biofilm but Far Behind Bacterial Biofilms

“…Interestingly, Galectin-3 and Tetherin, have been found in viral biofilm and could be involved in the retaining molecules early steps ( Pais-Correia et al, 2010 ; Nakamura et al, 2019 ). The viral biofilm is also composed of ECM components, like Agrin, HSPG, and Collagen IV (COL4A1 and COL4A2; Pais-Correia et al, 2010 ; Gross and Thoma-Kress, 2016 ; Millen et al, 2019 ). Moreover, Tarasevich et al showed that the cell surface proteins CD4, CD150, CD70, CD80, and CD25 are also present in viral biofilm structures ( Tarasevich et al, 2015 ).…”

“…It is for example, the case of Fucosyltransferase Fuc-T VII, which synthetizes sialyl Lewis X, but also of collagen IV (COL4A1, COL4A2; Hiraiwa et al, 1997 , 2003 ; Pais-Correia et al, 2010 ) and of ICAM-1 ( Hiraiwa et al, 2003 ; Fazio et al, 2019 ). Using transcriptome analysis, Millen et al showed recently that col4A1/2 genes are upregulated both in HTLV-1 infected cells and in Tax-induced cells, upon Tax-dependent transactivation of their promoter ( Millen et al, 2019 ). Overexpressed COL4 proteins co-localized with the viral Gag protein in viral biofilm and were involved in viral transmission.…”

Transfer of HTLV-1 p8 and Gag to target T-cells depends on VASP, a novel interaction partner of p8

“…To confirm these findings with different methods, we made use of our recently described flow cytometry-based assay [29] and generated stable Jurkat T-cells with a knockout of VASP applying CRISPR/Cas9 technology. For this purpose, we cloned two different VASP-specific guide RNAs into the lentiviral vector system lentiCRISPRv2 [53] and transduced Jurkat T-cells either with a pool of both lentiCRISPRv2-VASP-guide1 and lentiCRISPRv2-VASP-guide2 vectors or with the unspecific control vector lentiCRISPRv2-scramble-guide [19]. After transduction and selection, western blot analysis revealed that VASP protein was knocked out at day 14 and day 28 post transduction in the newly generated cell line Jurkat VASP-KO (knockout) compared to the Jurkat scramble control (S4A Fig).…”

“…For transmission at tight cell-cell contacts, two non-exclusive mechanisms of virus transmission at the virological synapse (VS), a virus-induced specialized cell-cell-contact, have been proposed: polarized budding of HTLV-1 into synaptic clefts [14] and cell surface transfer of viral biofilms [17]. For formation of the VS and the viral biofilm, the viral transactivator Tax plays a major role [18,19]. For transmission via cellular conduits, however, the accessory protein p8 is a prerequisite [15,16].…”