HLA-E: Strong association with β2-microglobulin and surface expression in the absence of HLA class I signal sequence-derived peptides

Elisa Lo Monaco, Leonardo Sibilio, Elisa Melucci, Elisa Tremante, Miloslav Suchànek, Vaclav Horejsi, Aline Martayan, Patrizio Giacomini

Research output: Contribution to journalArticle

Abstract

The nonclassical class I HLA-E molecule folds in the presence of peptide ligands donated by the signal sequences of permissive class I HLA alleles, with the aid of TAP and tapasin. To identify HLA-E-specific Abs, four monoclonals of the previously described MEM series were screened by isoelectric focusing (IEF) blot and immunoprecipitation/IEF on >30 single-allele class I transfectants and HLA-homozygous B lymphoid cells coexpressing HLA-E and HLA-A, -B, -C, -F, or -G. Despite their HLA-E-restricted reactivity patterns (MEM-E/02 in IEF blot; MEM-E/07 and MEM-E/08 in immunoprecipitation), all of the MEM Abs unexpectedly reacted with β2-microglobulin (β2m)-free and denatured (but not α2m-associated and folded) HLA-E H chains. Remarkably, other HLA-E-restricted Abs were also reactive with free H chains. Immunodepletion, in vitro assembly, flow cytometry, and three distinct surface-labeling methods, including a modified (conformation-independent) biotin-labeling assay, revealed the coexistence of HLA-E conformers with unusual and drastically antithetic features. MEM-reactive conformers were thermally unstable and poorly surface expressed, as expected, whereas β2m- associated conformers were either unstable and weakly reactive with the prototypic conformational Ab W6/32, or exceptionally stable and strongly reactive with Abs to β2m even in cells lacking permissive alleles (721.221), TAP (T2), or tapasin (721.220). Noncanonical, immature (endoglycosidase H-sensitive) HLA-E glycoforms were surface expressed in these cells, whereas mature glycoforms were exclusively expressed (and at much lower levels) in cells carrying permissive alleles. Thus, HLA-E is a good, and not a poor, β2m assembler, and TAP/tapasin-assisted ligand donation is only one, and possibly not even the major, pathway leading to its stabilization and surface expression.

Original languageEnglish
Pages (from-to)5442-5450
Number of pages9
JournalJournal of Immunology
Volume181
Issue number8
Publication statusPublished - 2008

ASJC Scopus subject areas

  • Immunology
  • Medicine(all)

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