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

PLoS Pathog. 2020 Sep 30;16(9):e1008879. doi: 10.1371/journal.ppat.1008879. eCollection 2020 Sep.

Abstract

The Human T-cell leukemia virus type 1 (HTLV-1) orf I-encoded accessory protein p8 is cleaved from its precursor p12, and both proteins contribute to viral persistence. p8 induces cellular protrusions, which are thought to facilitate transfer of p8 to target cells and virus transmission. Host factors interacting with p8 and mediating p8 transfer are unknown. Here, we report that vasodilator-stimulated phosphoprotein (VASP), which promotes actin filament elongation, is a novel interaction partner of p8 and important for p8 and HTLV-1 Gag cell-to-cell transfer. VASP contains an Ena/VASP homology 1 (EVH1) domain that targets the protein to focal adhesions. Bioinformatics identified a short stretch in p8 (amino acids (aa) 24-45) which may mediate interactions with the EVH1 domain of VASP. Co-immunoprecipitations confirmed interactions of VASP:p8 in 293T, Jurkat and HTLV-1-infected MT-2 cells. Co-precipitation of VASP:p8 could be significantly blocked by peptides mimicking aa 26-37 of p8. Mutational studies revealed that the EVH1-domain of VASP is necessary, but not sufficient for the interaction with p8. Further, deletion of the VASP G- and F-actin binding domains significantly diminished co-precipitation of p8. Imaging identified areas of partial co-localization of VASP with p8 at the plasma membrane and in protrusive structures, which was confirmed by proximity ligation assays. Co-culture experiments revealed that p8 is transferred between Jurkat T-cells via VASP-containing conduits. Imaging and flow cytometry revealed that repression of both endogenous and overexpressed VASP by RNA interference or by CRISPR/Cas9 reduced p8 transfer to the cell surface and to target Jurkat T-cells. Stable repression of VASP by RNA interference in chronically infected MT-2 cells impaired both p8 and HTLV-1 Gag transfer to target Jurkat T-cells, while virus release was unaffected. Thus, we identified VASP as a novel interaction partner of p8, which is important for transfer of HTLV-1 p8 and Gag to target T-cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Cell Adhesion Molecules* / chemistry
  • Cell Adhesion Molecules* / genetics
  • Cell Adhesion Molecules* / metabolism
  • Focal Adhesions* / chemistry
  • Focal Adhesions* / genetics
  • Focal Adhesions* / metabolism
  • Focal Adhesions* / virology
  • Gene Products, gag* / chemistry
  • Gene Products, gag* / genetics
  • Gene Products, gag* / metabolism
  • Human T-lymphotropic virus 1* / chemistry
  • Human T-lymphotropic virus 1* / genetics
  • Human T-lymphotropic virus 1* / metabolism
  • Humans
  • Jurkat Cells
  • Microfilament Proteins* / chemistry
  • Microfilament Proteins* / genetics
  • Microfilament Proteins* / metabolism
  • Phosphoproteins* / chemistry
  • Phosphoproteins* / genetics
  • Phosphoproteins* / metabolism
  • Protein Domains
  • T-Lymphocytes* / chemistry
  • T-Lymphocytes* / metabolism
  • T-Lymphocytes* / virology

Substances

  • Cell Adhesion Molecules
  • Gene Products, gag
  • Microfilament Proteins
  • Phosphoproteins
  • vasodilator-stimulated phosphoprotein

Grant support

This work was funded by the DFG (https://www.dfg.de/; grants TH 2166/1-1 and SFB796, C6 for AKTK, and SFB796, A2 for HS; GRK2504 A2 to AKTK and C2 to HS) and by the Interdisciplinary Center for Clinical Research at the Medical Faculty of the Friedrich-Alexander-Universität (FAU) Erlangen-Nürnberg (https://www.izkf.med.fau.de/; project A91, AKTK). AKTK received also funding from the ELAN funds at the University Hospital of the FAU Erlangen-Nuremberg (https://www.izkf.med.fau.de/elan/). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. We acknowledge support by Deutsche Forschungsgemeinschaft and Friedrich-Alexander-Universität Erlangen-Nürnberg (FAU) within the funding program Open Access Publishing.