Molecular mapping of signals in the Qa-2 antigen required for attachment of the phosphatidylinositol membrane anchor.
Proteins anchored in the membrane by covalent linkage to phosphatidylinositol (PtdIns) can be released by treatment with purified PtdIns-specific phospholipase C (Ptd-Ins-PLC). A recent survey of leukocyte antigens using flow cytometry has shown that staining of certain Qa antigens was diminished after PtdIns-PLC treatment, but staining of structurally related H-2 antigens was not affected. Therefore, in this study, the sensitivity of cell-surface Qa-2, H-2Kb, and H-2Db to hydrolysis by PtdIns-PLC was investigated biochemically by immunoprecipitation of radioiodinated molecules from cell lysates or supernatants. Qa-2, but not H-2Kb, was released from the surface of PtdIns-PLC-treated C57BL/10 mouse spleen cells and recovered in the cell supernatants. Similar analysis of thymoma cells transfected with cloned C57BL/10 genes showed that cell-surface Qa-2 molecules encoded by a Q7b cDNA and the Q7b or Q9b gene were sensitive to hydrolysis by PtdIns-PLC, whereas the H-2Kb and H-2Db gene products were resistant. Using thymoma cells transfected with hybrid genes constructed by exchanging exons between Q7b and H-2Db, the signals for PtdIns modification were localized to a defined region of Qa-2. This region differs from H-2Db most significantly by the presence of a central aspartate residue in the transmembrane segment and in the length of the cytoplasmic portion.
Waneck
G L
GL
Biogen Research Corporation, Cambridge, MA 02142.
Sherman
D H
DH
Kincade
P W
PW
Low
M G
MG
Flavell
R A
RA
D000595:Amino Acid Sequence; D000818:Animals; D000954:Antigens, Surface / Q000235:genetics* / Q000276:immunology; D002460:Cell Line; D005796:Genes; D006183:H-2 Antigens / Q000235:genetics* / Q000276:immunology; D006649:Histocompatibility Antigens / Q000235:genetics* / Q000276:immunology; D015395:Histocompatibility Antigens Class I; D008563:Membrane Lipids / Q000276:immunology*; D051379:Mice; D008969:Molecular Sequence Data; D010716:Phosphatidylinositols / Q000276:immunology*
D000954:Antigens, Surface; D006183:H-2 Antigens; D006649:Histocompatibility Antigens; D015395:Histocompatibility Antigens Class I; D008563:Membrane Lipids; D010716:Phosphatidylinositols; C032907:Q surface antigens