Impaired assembly results in the accumulation of multiple HLA-C heavy chain folding intermediates

Leonardo Sibilio, Aline Martayan, Andrea Setini, Rocco Fraioli, Doriana Fruci, Jeffrey Shabanowitz, Donald F. Hunt, Patrizio Giacomini

Research output: Contribution to journalArticlepeer-review

Abstract

Class I MHC H chains assemble with β2-microglobulin (β2m) and are loaded with peptide Ags through multiple foiding steps. When free of β2m, human H chains react with Abs to linear epitopes, such as L31. Immunodepletion and coimmunoprecipitation experiments, performed in tins study, detected a preferential association of L31-reactive, β2m-free H chains with calnexin in β2m-defective cells, and with calreticulin and TAP in β2m-expressing cells. In β2m-defective cells, the accumulation of calnexin-bound H chains stoichiometrically exceeded their overall accumulation, a finding that supports both chaperoning preferences and distinct sorting abilities for different class I folds. No peptide species, in a mass range compatible with that of the classical class I Iigands, could be detected by mass spectrometry of acidic ehiates from L31-reactive HLA-Cw1 H chains. In vitro assembly experiments in TAP-defective T2 cells, and in cells expressing an intact Ag-processing machinery, demonstrated that L31 H chains are not only free of, but also unreceptive to, peptides. L31 and HC10, which bind nearly adjacent linear epitopes of the α1 domain α helix, reciprocally immunodepleted free HLA-C H chains, indicating the existence of a local un-/mis-folding involving the N-terminal end of the α domain α helix and peptide-anchoring residues of the class I H chain. Thus, unlike certain murine free H chains, L31-reactive H chains are not the immediate precursors of conformed class I molecules. A model inferring their precursor-product relationships with other known class I intermediates is presented.

Original languageEnglish
Pages (from-to)6651-6658
Number of pages8
JournalJournal of Immunology
Volume175
Issue number10
Publication statusPublished - Nov 15 2005

ASJC Scopus subject areas

  • Immunology

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