Degradation of T-cell receptor chains in the endoplasmic reticulum is inhibited by inhibitors of cysteine proteases.
The endoplasmic reticulum, or an organelle closely associated with it, contains proteases that can be used to remove partially assembled or improperly folded proteins. Very little is known at present about the types of protease that degrade these proteins. The beta chain and cluster of differentiation (CD)3 delta subunit of the human T-cell antigen receptor (TCR) are degraded shortly after synthesis. In this study Chinese hamster ovary (CHO) cells transfected with either beta or delta were incubated with a panel of protease inhibitors, and the rates of degradation of the transfected proteins were followed using chain-specific enzyme-linked immunosorbent assays (ELISAs). Of the protease inhibitors tested, degradation of both chains was highly sensitive to sulfhydryl reagents and peptidyl inhibitors of cysteine proteases. Concentrations of inhibitors that produced near complete inhibition of degradation in the endoplasmic reticulum did not cause gross changes in cellular ATP levels nor did they significantly slow constitutive secretion from CHO cells. The inhibitors did not affect the ability of CHO cells to synthesize and assemble disulphide-linked TCR zeta dimers. We conclude that the protease inhibitors were not toxic to cells and did not affect the biosynthetic activity of the endoplasmic reticulum. Furthermore, they did not alter the ability of the endoplasmic reticulum to deliver its content to the Golgi apparatus. Taken together, these results suggest that the cysteine protease inhibitors slow degradation in the endoplasmic reticulum through an action on cysteine proteases. The results imply that the endoplasmic reticulum contains cysteine proteases that can be used to remove retained proteins.
Wileman
T
T
Dana Farber Cancer Institute, Department of Pathology, Harvard Medical School, Boston, Massachusetts 02115.
Kane
L P
LP
Terhorst
C
C
D000255:Adenosine Triphosphate / Q000378:metabolism; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003513:Cycloheximide / Q000494:pharmacology; D003546:Cysteine Endopeptidases / Q000494:pharmacology*; D004591:Electrophoresis, Polyacrylamide Gel; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D004797:Enzyme-Linked Immunosorbent Assay; D006023:Glycoproteins / Q000494:pharmacology; D007460:Iodoacetamide / Q000494:pharmacology; D016693:Receptors, Antigen, T-Cell, alpha-beta / Q000378:metabolism*; D016692:Receptors, Antigen, T-Cell, gamma-delta / Q000378:metabolism*; D014108:Tosylphenylalanyl Chloromethyl Ketone / Q000494:pharmacology
D000590:Amino Acid Chloromethyl Ketones; D006023:Glycoproteins; D016693:Receptors, Antigen, T-Cell, alpha-beta; D016692:Receptors, Antigen, T-Cell, gamma-delta; C039304:calpain inhibitors; C006735:ZPCK; D014108:Tosylphenylalanyl Chloromethyl Ketone; D000255:Adenosine Triphosphate; D003513:Cycloheximide; D003546:Cysteine Endopeptidases; D007460:Iodoacetamide