Calnexin Association Is Not Sufficient to Protect T Cell Receptor α Proteins from Rapid Degradation in CD4+CD8+ Thymocytes

doi:10.1074/jbc.273.37.23674

Calnexin Association Is Not Sufficient to Protect T Cell Receptor α Proteins from Rapid Degradation in CD4⁺CD8⁺ Thymocytes*

From the Department of Microbiology & Immunology, East Carolina University, School of Medicine, Greenville, North Carolina 27858-4354 and ^‡Experimental Immunology Branch, NCI, National Institutes of Health, Bethesda, Maryland 20892-1360

Abstract

During T cell development, assembly of the mutisubunit T cell receptor (TCR) complex is regulated by the differential stability of newly synthesized TCRα molecules, having a half-life of approximately 20 min in immature CD4⁺CD8⁺ thymocytes compared with >75 min in mature T cells. The molecular basis for TCRα instability in CD4⁺CD8⁺ thymocytes is unknown but has been postulated to involve abnormalities in N-glycan processing and calnexin assembly as perturbation of these pathways markedly destabilizes TCRα proteins in all other T cell types examined. Here, we compared the processing of TCRα glycoproteins and their assembly with calnexin and calreticulin chaperones in CD4⁺CD8⁺ thymocytes and splenic T cells. These studies show that TCRα glycoproteins synthesized in CD4⁺CD8⁺ thymocytes were processed in a similar manner as those made in splenic T cells and that TCRα proteins stably associated with calnexin in both cell types. Interestingly, however, TCRα association with the calnexin-related molecule calreticulin was decreased in CD4⁺CD8⁺thymocytes compared with splenic T cells. Finally, TCRα degradation in CD4⁺CD8⁺ thymocytes was impaired by inhibitors of proteasome activity, which was correlated with stabilization of calnexin·TCRα complexes. These data demonstrate that calnexin association is not sufficient to protect TCRα proteins from rapid degradation in CD4⁺CD8⁺ thymocytes, suggesting that additional components of the quality control system of the endoplasmic reticulum operate to ensure the proper folding of nascent TCRα glycoproteins.

Footnotes

↵* This work was supported by a faculty development research grant from the East Carolina University School of Medicine and National Institutes of Health Grant R29 AI42104 (to K. P. K.).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked “advertisement” in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
↵§ Current address: Cell Biology and Metabolism Branch, NICHD, National Institutes of Health, Bethesda, MD 20892-5430.
↵¶ To whom correspondence should be addressed. Tel.: 252-816-2703; Fax: 252-816-3104; E-mail:kearse{at}brody.med.ecu.edu.
Abbreviations:

ER

endoplasmic reticulum

TCR

T cell receptor

Ab

antibody

mAb

monoclonal Ab

TPCK

N-tosyl-l-phenylalanine chloromethyl ketone

Endo H

endoglycosidase H

PAGE

polyacrylamide gel electrophoresis

Glc

glucose

cas

castanospermine.
- Received March 18, 1998.
- Revision received May 8, 1998.
The American Society for Biochemistry and Molecular Biology, Inc.