Role of ESCRT-I in retroviral budding

J Virol. 2003 Apr;77(8):4794-804. doi: 10.1128/jvi.77.8.4794-4804.2003.

Abstract

Retroviral late-budding (L) domains are required for the efficient release of nascent virions. The three known types of L domain, designated according to essential tetrapeptide motifs (PTAP, PPXY, or YPDL), each bind distinct cellular cofactors. We and others have demonstrated that recruitment of an ESCRT-I subunit, Tsg101, a component of the class E vacuolar protein sorting (VPS) machinery, is required for the budding of viruses, such as human immunodeficiency virus type 1 (HIV-1) and Ebola virus, that encode a PTAP-type L domain, but subsequent events remain undefined. In this study, we demonstrate that VPS28, a second component of ESCRT-I, binds to a sequence close to the Tsg101 C terminus and is therefore recruited to the plasma membrane by HIV-1 Gag. In addition, we show that Tsg101 exhibits a multimerization activity. Using a complementation assay in which Tsg101 is artificially recruited to sites of HIV-1 assembly, we demonstrate that the integrity of the VPS28 binding site within Tsg101 is required for particle budding. In addition, mutation of a putative leucine zipper or residues important for Tsg101 multimerization also impairs the ability of Tsg101 to support HIV-1 budding. A minimal multimerizing Tsg101 domain is a dominant negative inhibitor of PTAP-mediated HIV-1 budding but does not inhibit YPDL-type or PPXY-type L-domain function. Nevertheless, YDPL-type L-domain activity is inhibited by expression of a catalytically inactive mutant of the class E VPS ATPase VPS4. These results indicate that all three classes of retroviral L domains require a functioning class E VPS pathway in order to effect budding. However, the PTAP-type L domain appears to be unique in its requirement for an intact, or nearly intact, ESCRT-I complex.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Binding Sites
  • Carrier Proteins / chemistry
  • Carrier Proteins / genetics
  • Carrier Proteins / metabolism
  • Cell Line
  • DNA-Binding Proteins / chemistry
  • DNA-Binding Proteins / genetics
  • DNA-Binding Proteins / metabolism*
  • Dimerization
  • Endosomal Sorting Complexes Required for Transport
  • HIV-1 / genetics
  • HIV-1 / growth & development*
  • HIV-1 / pathogenicity
  • Humans
  • Leucine Zippers / genetics
  • Leukemia Virus, Murine / genetics
  • Leukemia Virus, Murine / growth & development*
  • Leukemia Virus, Murine / pathogenicity
  • Mice
  • Molecular Sequence Data
  • Saccharomyces cerevisiae Proteins*
  • Transcription Factors / chemistry
  • Transcription Factors / genetics
  • Transcription Factors / metabolism*
  • Two-Hybrid System Techniques
  • Vesicular Transport Proteins*
  • Virion / metabolism

Substances

  • Carrier Proteins
  • DNA-Binding Proteins
  • Endosomal Sorting Complexes Required for Transport
  • Saccharomyces cerevisiae Proteins
  • Transcription Factors
  • Tsg101 protein
  • VPS28 protein, S cerevisiae
  • Vesicular Transport Proteins