The Inner Loop of Tetraspanins CD82 and CD81 Mediates Interactions with Human T Cell Lymphotrophic Virus Type 1 Gag Protein

Gisela Heidecker,Dmitriy Mazurov,David Derse

doi:10.1074/jbc.m607322200

Gisela Heidecker, Dmitriy Mazurov + Show 1 more

Open Access

https://doi.org/10.1074/jbc.m607322200

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Abstract

The tetraspanin superfamily proteins play important roles in organizing membrane protein complexes, modulating integrin function, and controlling T cell adhesion. Tetraspanins such as CD82 contain two extracellular loops with its N terminus, C terminus, and inner loop exposed to the cytoplasm. The matrix (MA) domain of human T cell lymphotrophic virus, type 1 (HTLV-1), Gag interacts with the cytoplasmic face of the plasma membrane and is concentrated at tetraspanin-enriched microdomains. To understand the basis of this association, we generated site-directed mutations in the various domains of CD82 and used coimmunoprecipitation and colocalization approaches to examine interactions with HTLV-1 MA. The large extracellular loop of CD82, which is important for interactions with integrins, was not required for the association with HTLV-1 MA. The cytoplasmic N terminus and C terminus of CD82 were also dispensable for CD82-MA interactions. In contrast, mutations of conserved amino acids in the inner loop of CD82 or of palmitoylated cysteines that flank the inner loop diminished CD82 association with MA. HTLV-1 MA also interacted with the inner loop of CD81. Thus, association of HTLV-1 Gag with tetraspanin-enriched microdomains is mediated by the inner loops of CD81 and CD82.

Highlights

The tetraspanins are a large family of four transmembrane domain-containing proteins that are widely expressed in many cell types and tissues
We previously showed that HTLV-1 Gag localizes to tetraspanin-enriched microdomains in the plasma membrane and that a dimer of HTLV-1 MA coimmunoprecipitates with CD82 from cell extracts prepared in extraction buffers containing Brij97 detergent [25]
Human 293T cells were cotransfected with an HTLV-1 Gag expression vector in combination with either wild type or SSG CD82 expression plasmids in which a FLAG epitope tag was appended to the N terminus of CD82

Summary

Introduction

The tetraspanins are a large family of four transmembrane domain-containing proteins that are widely expressed in many cell types and tissues. If HTLV-1 Gag associates directly with CD82, it is likely to interact with a region of the protein exposed to the cytoplasm (Fig. 1, top), because Gag is an intracellular protein, anchored to the inner leaflet of the plasma membrane by myristate modification and specific amino acids in the MA domain [31, 32].

Results

Conclusion