Distinct MHC Class II Molecules Are Associated on the Dendritic Cell Surface in Cholesterol-dependent Membrane Microdomains

Sanjay Khandelwal,Paul A Roche

doi:10.1074/jbc.m110.147793

Sanjay Khandelwal, Paul A Roche

Open Access

https://doi.org/10.1074/jbc.m110.147793

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Abstract

Very small amounts of MHC class II-peptide complexes expressed on the surface of antigen-presenting cells (APCs) are capable of stimulating antigen-specific CD4 T cells. There is intense interest to elucidate the molecular mechanisms by which these small amounts of MHC-II can cluster, cross-link T cell receptors, and promote T cell proliferation. We now demonstrate that a significant fraction of the total pool of MHC-II molecules on the surface of dendritic cells is physically associated in macromolecular aggregates. These MHC-II/MHC-II interactions have been probed by co-immunoprecipitation analysis of the MHC-II I-A molecule with the related I-E molecule. These molecular associations are maintained in gentle detergents but are disrupted in harsh detergents such as Triton X-100. MHC-II I-A/I-E interactions are disrupted when plasma membrane cholesterol is extracted using methyl β-cyclodextrin, suggesting that lipid raft microdomains are important mediators of these MHC-II interactions. Although it has been proposed that tetraspanin proteins regulate molecular clustering, aggregation, and co-immunoprecipitation in APCs, genetic deletion of the tetraspanin family members CD9 or CD81 had no effect on MHC-II I-A/I-E binding. These data demonstrate that the presence of distinct forms of MHC-II with plasma membrane lipid rafts is required for MHC-II aggregation in APCs and provides a molecular mechanism allowing dendritic cells expressing small amounts of MHC-II-peptide complexes to cross-link and stimulate CD4 T cells.

Highlights

MHC class II molecules expressed on the surface of specialized antigen presenting cells (APCs)2 function by presenting peptides derived from foreign protein antigens to specific CD4 T cells, a process that initiates and propagates immune responses [1]
I-A and I-E Molecules Associate on the Plasma Membrane of dendritic cells (DCs)—Cells lysed in gentle detergents such as Brij-58 maintain membrane microdomain associations, whereas those lysed under more harsh detergent conditions, such as in Triton X-100, do not [33,34,35]
CD4 T cell activation requires the cross-linking of antigenspecific T cell receptors for antigen (TCR) by relevant MHC-II-peptide complexes expressed on antigen-presenting cells (APCs) [42]

Summary

Introduction

MHC class II molecules expressed on the surface of specialized antigen presenting cells (APCs) function by presenting peptides derived from foreign protein antigens to specific CD4 T cells, a process that initiates and propagates immune responses [1]. The interaction of MHC-II-peptide complexes on APCs with T cell receptors for antigen (TCR) on T cells cross-links the TCR, thereby initiating a signaling cascade that leads to cytokine production and T cell proliferation In addition to associating with lipid raft membrane microdomains, MHC-II binds to a class of proteins termed tetraspanins (19 –25). Tetraspanins form lateral associations with many different proteins, and tetraspan microdomains are important in a wide variety of processes, including signal transduction, cell proliferation, cell adhesion, cell migration, cell fusion, and host parasite interactions (26 –28). Most importantly, these proteins play a key role in the function of many immune cells. The tetraspanins CD9 and CD81 can provide a “co-stimulatory-like”

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