Peptide and beta 2-microglobulin regulation of cell surface MHC class I conformation and expression.
We have examined the roles of peptide and beta 2-microglobulin (beta 2m) in regulating the conformation and expression level of class I molecules on the cell surface. Using a cell line synthesizing H-2Dd H chain and mouse beta 2m but defective in endogenous peptide loading, we demonstrate the ability of either exogenous peptide or beta 2m alone to increase surface H-2Dd expression at both 25 degrees C and 37 degrees C. Peptide and beta 2m show marked synergy in their abilities to increase surface class I expression, with minimal increases promoted by peptide in the absence of free beta 2m. Low temperature-induced molecules have indistinguishable rates of loss of beta 2m and alpha 1/alpha 2 domain conformational epitopes during culture at 37 degrees C. However, the rate of alpha 3 epitope loss is much slower, indicating a minimum of two steps in class I loss from the cell surface: 1) loss of beta 2m binding to H chain and unfolding of the alpha 1/alpha 2 region; then 2) denaturation, degradation, or internalization of the free H chains possessing alpha 3 epitopes. These data show for the first time that free H chains survive for a finite time on the membrane in a form capable of refolding into alpha 1/alpha 2 epitope positive molecules upon addition of beta 2m and peptide. This refolding in the presence of beta 2m and peptide can explain the reported requirement for both components in sensitizing cells for class I-dependent CTL lysis. It also indicates that such conformational changes in class I molecules are not strictly dependent on either newly synthesized H chains or on intracellular chaperons. The study of H chain-peptide-beta 2m interaction on the cell surface may be relevant to understanding intracellular peptide loading events.
Otten
G R
GR
Lymphocyte Biology Section, National Institute of Allergy and Infectious Disease, National Institutes of Health, Bethesda, MD 20892.
Bikoff
E
E
Ribaudo
R K
RK
Kozlowski
S
S
Margulies
D H
DH
Germain
R N
RN
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001692:Biological Transport; D002451:Cell Compartmentation; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D006183:H-2 Antigens / Q000378:metabolism* / Q000648:ultrastructure; D066298:In Vitro Techniques; D008565:Membrane Proteins / Q000378:metabolism / Q000648:ultrastructure; D051379:Mice; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry* / Q000378:metabolism; D011487:Protein Conformation; D001613:beta 2-Microglobulin / Q000502:physiology*
D000911:Antibodies, Monoclonal; D006183:H-2 Antigens; D008565:Membrane Proteins; D010455:Peptides; D001613:beta 2-Microglobulin