Granzyme B-mediated degradation of T-cell receptor zeta chain.
We recently reported that the T-cell receptor (TCR)-zeta chain is cleaved by caspase-3 and -7 in apoptotic T lymphocytes or in a cell-free system. We report here that the zeta chain is also a direct substrate for granzyme B (GrB) proteolytic activity. Loss in expression of TCR-zeta was observed in Jurkat T leukemic cells treated by a combination of GrB and a replication-deficient adenovirus. Although the apoptosis initiated in these cells by GrB was significantly reduced by the pancaspase inhibitor Z-VAD-FMK, TCR-zeta degradation was not prevented. These findings suggest that the GrB-mediated degradation of TCR-zeta chain can proceed despite the efficient inhibition of caspase activity. An in vitro translated TCR-zeta product was efficiently cleaved by GrB, which suggests that the TCR-zeta protein is a direct substrate for GrB. As assessed by site-directed mutagenesis, the activity of GrB was directed toward aspartic acid residues that were different from those of recombinant caspase-3. Whereas caspase-3 cleavage products appear to accumulate, the GrB-generated products seem to undergo further degradation, which suggests the presence of multiple GrB-specific cleavage sites within the TCR-zeta protein. These findings suggest that the TCR-zeta protein in target T lymphocytes serves as a substrate for the proteolytic activities that are featured by the two major mechanisms of cytotoxicity: death receptor pathways mediated by caspases and granule exocytosis mediated by direct GrB activity or GrB-activated caspases. TCR-zeta protein degradation may be of significance in cytotoxic mechanisms directed against T cells infected with viruses, such as HIV-1, in which the TCR-zeta protein is used for viral pathogenesis.
Department of Pathology, The University of Pittsburgh School of Medicine, Pennsylvania 15213, USA.
Eva
E
Wieckowski
Gui-Qiang
GQ
Wang
Brian R
BR
Gastman
Leslie A
LA
Goldstein
Hannah
H
Rabinowich
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology; D053148:Caspase 3; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D053804:Granzymes; D006801:Humans; D019169:Jurkat Cells / Q000187:drug effects / Q000378:metabolism / Q000473:pathology; D008565:Membrane Proteins / Q000378:metabolism*; D011948:Receptors, Antigen, T-Cell / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000378:metabolism / Q000494:pharmacology*
D000590:Amino Acid Chloromethyl Ketones; D061945:Caspase Inhibitors; D015853:Cysteine Proteinase Inhibitors; D008565:Membrane Proteins; D011948:Receptors, Antigen, T-Cell; C054979:antigen T cell receptor, zeta chain; C096713:benzyloxycarbonylvalyl-alanyl-aspartyl fluoromethyl ketone; C509213:GZMB protein, human; D053804:Granzymes; D012697:Serine Endopeptidases; C505407:CASP3 protein, human; D053148:Caspase 3; D020169:Caspases