Processing of a viral glycoprotein in the endoplasmic reticulum for class II presentation.
Endogenous processing of viral glycoproteins for presentation to CD4+T cells is a poorly investigated aspect of antigen processing and presentation. This pathway may involve not only pathogens, but also self proteins, and may thus be involved in self-tolerance. We have characterized the processing of the endoplasmic reticulum-restricted glycoprotein (G) of vesicular stomatitis virus, termed poison tail (Gpt), biochemically and enzymatically, and by T cell recognition assays. Expressed with a vaccinia vector, Gpt remains endoglycosidase H-sensitive and does not mature to endoglycosidase D sensitivity. The protein is degraded in the ER with a T1/2 of 4 h. Gpt peptides are not secreted since Gpt-infected cells are unable to sensitize uninfected antigen-presenting cells in an innocent bystander assay. Using flow cytometry, Gpt is undetectable on the plasma membrane; in contrast, wild-type G is readily found on the surface or secreted into the milieu as soluble G following infection of A20 cells with a vaccinia recombinant expressing G. The degradation of Gpt is sensitive to the thiol reagent diamide and occurs optimally at physiological pH. A series of proteolytic inhibitors were tested: 3,4-dichloroisocoumarin and 1-chloro-3-tosylamido-7-amino-2-heptanone inhibited degradation, which suggests the involvement of a serine protease. The degradation does not require transport to the Golgi complex, and is not sensitive to a variety of lysosomotropic agents. We show that the degradation products include the immunogenic epitopes recognized by a panel of T cell clones and hybridomas.
Bartido
S M
SM
Biology Department, New York University, New York 10003-6688, USA.
Diment
S
S
Reiss
C S
CS
D000818:Animals; D017951:Antigen Presentation / Q000276:immunology*; D015496:CD4-Positive T-Lymphocytes / Q000276:immunology; D016466:CHO Cells; D002460:Cell Line; D006224:Cricetinae; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006023:Glycoproteins / Q000276:immunology / Q000378:metabolism; D006056:Golgi Apparatus / Q000187:drug effects / Q000378:metabolism; D000882:Haplorhini; D000949:Histocompatibility Antigens Class II / Q000235:genetics*; D006863:Hydrogen-Ion Concentration; D051379:Mice; D008807:Mice, Inbred BALB C; D013438:Sulfhydryl Compounds / Q000494:pharmacology; D014721:Vesicular stomatitis Indiana virus / Q000276:immunology*; D014759:Viral Envelope Proteins / Q000276:immunology* / Q000378:metabolism*
D006023:Glycoproteins; D000949:Histocompatibility Antigens Class II; D013438:Sulfhydryl Compounds; D014759:Viral Envelope Proteins