FUNCTIONAL CELL-SURFACE EXPRESSION BY A RECOMBINANT SINGLE-CHAIN CLASS-I MAJOR HISTOCOMPATIBILITY COMPLEX MOLECULE WITH A CIS-ACTIVE BETA(2)-MICROGLOBULIN DOMAIN

被引:20
作者
LEE, L
MCHUGH, L
RIBAUDO, RK
KOZLOWSKI, S
MARGULIES, DH
MAGE, MG
机构
[1] NCI,BIOCHEM LAB,BETHESDA,MD 20892
[2] NIAID,IMMUNOL LAB,BETHESDA,MD 20892
关键词
MAJOR HISTOCOMPATIBILITY COMPLEX CLASS I; SINGLE CHAIN; BETA(2)-MICROGLOBULIN;
D O I
10.1002/eji.1830241110
中图分类号
R392 [医学免疫学]; Q939.91 [免疫学];
学科分类号
100102 ;
摘要
As a preliminary step towards the use of cell surface single-chain class I major histocompatibility complex (MHC) molecules as T cell immunogens, we have engineered a recombinant gene encoding a full-length cell surface single-chain version of the H-2D(d) class I MHC molecule (SC beta D(d)m) which has beta(2)-microglobulin (beta(2)m) covalently linked to the amino terminus of a full-length H-2D(d) heavy chain via a peptide spacer. The single-chain protein is correctly folded and stably expressed on the surface of transfected L cells. It can present an antigenic peptide to an H-2D(d)-restricted antigen-specific T cell hybridoma. When expressed in peptide-transport-deficient cells, SC beta D(d)m can be stabilized and pulsed for antigen presentation by incubation with extracellular peptide at 27 degrees or 37 degrees C, allowing the preparation of cells with single-chain molecules that are loaded with a single chosen antigenic peptide. SC beta D(d)m can be stably expressed in beta(2)m-negative cells, showing that the single-chain molecule uses its own beta(2)m domain to achieve correct folding and surface expression. Furthermore, the beta(2)m domain of SC beta D(d)m, unlike transfected free beta(2)m, does not rescue surface expression of endogenous class I MHC in the beta(2)m-negative cells. This strict cis activity of the beta(2)m domain of SC beta D(d)m makes possible the investigation of class I MHC function in cells, and potentially in animals, that express but a single type of class I MHC molecule.
引用
收藏
页码:2633 / 2639
页数:7
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