Initiation of Apoptosis by Granzyme B Requires Direct Cleavage of Bid, but Not Direct Granzyme B-mediated Caspase Activation

Overview

Journal J Exp Med

Specialties Allergy & Immunology
General Medicine

Date 2000 Nov 21

PMID 11085743

Citations 108

Authors

V R Sutton

J E Davis

M Cancilla

R W Johnstone

A A Ruefli

K Sedelies

K A Browne

J A Trapani

Affiliations

Soon will be listed here.

Abstract

The essential upstream steps in granzyme B-mediated apoptosis remain undefined. Herein, we show that granzyme B triggers the mitochondrial apoptotic pathway through direct cleavage of Bid; however, cleavage of procaspases was stalled when mitochondrial disruption was blocked by Bcl-2. The sensitivity of granzyme B-resistant Bcl-2-overexpressing FDC-P1 cells was restored by coexpression of wild-type Bid, or Bid with a mutation of its caspase-8 cleavage site, and both types of Bid were cleaved. However, Bid with a mutated granzyme B cleavage site remained intact and did not restore apoptosis. Bid with a mutation preventing its interaction with Bcl-2 was cleaved but also failed to restore apoptosis. Rapid Bid cleavage by granzyme B (<2 min) was not delayed by Bcl-2 overexpression. These results clearly placed Bid cleavage upstream of mitochondrial Bcl-2. In granzyme B-treated Jurkat cells, endogenous Bid cleavage and loss of mitochondrial membrane depolarization occurred despite caspase inactivation with z-Val-Ala-Asp-fluoromethylketone or Asp-Glu-Val-Asp-fluoromethylketone. Initial partial processing of procaspase-3 and -8 was observed irrespective of Bcl-2 overexpression; however, later processing was completely abolished by Bcl-2. Overall, our results indicate that mitochondrial perturbation by Bid is necessary to achieve a lethal threshold of caspase activity and cell death due to granzyme B.

Citing Articles

New Paradigms in Rejection Monitoring: Lymphocyte Subsets as Noninvasive Graft Markers in Vascularized Composite Allotransplantation.

Chinta S, Shah A, Tran D, Lee W, Mangiola M, Gelb B Plast Reconstr Surg Glob Open. 2025; 13(3):e6598.

PMID: 40051973 PMC: 11884835. DOI: 10.1097/GOX.0000000000006598.

The roles of CD4 T cell help, sex, and dose in the induction of protective CD8 T cells against a lethal poxvirus by mRNA-LNP vaccines.

Kafle S, Montoya B, Tang L, Tam Y, Muramatsu H, Pardi N Mol Ther Nucleic Acids. 2024; 35(3):102279.

PMID: 39188304 PMC: 11345529. DOI: 10.1016/j.omtn.2024.102279.

Natural Killer cells at the frontline in the fight against cancer.

Coenon L, Geindreau M, Ghiringhelli F, Villalba M, Bruchard M Cell Death Dis. 2024; 15(8):614.

PMID: 39179536 PMC: 11343846. DOI: 10.1038/s41419-024-06976-0.

Pharmacologic inhibition of dipeptidyl peptidase 1 (cathepsin C) does not block granzyme-mediated target cell killing by CD8 T or NK cells.

Sutton V, Watt S, Akhlaghi H, Cipolla D, Chen K, LaSala D Front Pharmacol. 2024; 15:1396710.

PMID: 39021839 PMC: 11251990. DOI: 10.3389/fphar.2024.1396710.

Granzymes in health and diseases: the good, the bad and the ugly.

Cigalotto L, Martinvalet D Front Immunol. 2024; 15():1371743.

PMID: 38646541 PMC: 11026543. DOI: 10.3389/fimmu.2024.1371743.

References

Vaux D, Haecker G, Strasser A . An evolutionary perspective on apoptosis. Cell. 1994; 76(5):777-9. DOI: 10.1016/0092-8674(94)90350-6. View

Fernandes-Alnemri T, Armstrong R, Krebs J, Srinivasula S, Wang L, Bullrich F . In vitro activation of CPP32 and Mch3 by Mch4, a novel human apoptotic cysteine protease containing two FADD-like domains. Proc Natl Acad Sci U S A. 1996; 93(15):7464-9. PMC: 38767. DOI: 10.1073/pnas.93.15.7464. View

Verhagen A, Ekert P, Pakusch M, Silke J, Connolly L, Reid G . Identification of DIABLO, a mammalian protein that promotes apoptosis by binding to and antagonizing IAP proteins. Cell. 2000; 102(1):43-53. DOI: 10.1016/s0092-8674(00)00009-x. View

Du C, Fang M, Li Y, Li L, Wang X . Smac, a mitochondrial protein that promotes cytochrome c-dependent caspase activation by eliminating IAP inhibition. Cell. 2000; 102(1):33-42. DOI: 10.1016/s0092-8674(00)00008-8. View

Schmitz I, Walczak H, Krammer P, Peter M . Differences between CD95 type I and II cells detected with the CD95 ligand. Cell Death Differ. 2000; 6(9):821-2. DOI: 10.1038/sj.cdd.4400569. View

Bruey J, Ducasse C, Bonniaud P, Ravagnan L, Susin S, Diaz-Latoud C . Hsp27 negatively regulates cell death by interacting with cytochrome c. Nat Cell Biol. 2000; 2(9):645-52. DOI: 10.1038/35023595. View

Bossy-Wetzel E, Newmeyer D, Green D . Mitochondrial cytochrome c release in apoptosis occurs upstream of DEVD-specific caspase activation and independently of mitochondrial transmembrane depolarization. EMBO J. 1998; 17(1):37-49. PMC: 1170356. DOI: 10.1093/emboj/17.1.37. View

Andrade F, Roy S, Nicholson D, Thornberry N, Rosen A, Casciola-Rosen L . Granzyme B directly and efficiently cleaves several downstream caspase substrates: implications for CTL-induced apoptosis. Immunity. 1998; 8(4):451-60. DOI: 10.1016/s1074-7613(00)80550-6. View

Kagi D, Ledermann B, Burki K, Seiler P, Odermatt B, Olsen K . Cytotoxicity mediated by T cells and natural killer cells is greatly impaired in perforin-deficient mice. Nature. 1994; 369(6475):31-7. DOI: 10.1038/369031a0. View

10.

Boldin M, Goncharov T, Goltsev Y, Wallach D . Involvement of MACH, a novel MORT1/FADD-interacting protease, in Fas/APO-1- and TNF receptor-induced cell death. Cell. 1996; 85(6):803-15. DOI: 10.1016/s0092-8674(00)81265-9. View

11.

Muzio M, Chinnaiyan A, Kischkel F, ORourke K, Shevchenko A, Ni J . FLICE, a novel FADD-homologous ICE/CED-3-like protease, is recruited to the CD95 (Fas/APO-1) death--inducing signaling complex. Cell. 1996; 85(6):817-27. DOI: 10.1016/s0092-8674(00)81266-0. View

12.

Yang X, Stennicke H, Wang B, Green D, Janicke R, Srinivasan A . Granzyme B mimics apical caspases. Description of a unified pathway for trans-activation of executioner caspase-3 and -7. J Biol Chem. 1998; 273(51):34278-83. DOI: 10.1074/jbc.273.51.34278. View

13.

Trapani J, Browne K, Smyth M, Jans D . Localization of granzyme B in the nucleus. A putative role in the mechanism of cytotoxic lymphocyte-mediated apoptosis. J Biol Chem. 1996; 271(8):4127-33. DOI: 10.1074/jbc.271.8.4127. View

14.

Srinivasula S, Fernandes-Alnemri T, Zangrilli J, Robertson N, Armstrong R, Wang L . The Ced-3/interleukin 1beta converting enzyme-like homolog Mch6 and the lamin-cleaving enzyme Mch2alpha are substrates for the apoptotic mediator CPP32. J Biol Chem. 1996; 271(43):27099-106. DOI: 10.1074/jbc.271.43.27099. View

15.

Harvey N, Trapani J, Fernandes-Alnemri T, Litwack G, Alnemri E, Kumar S . Processing of the Nedd2 precursor by ICE-like proteases and granzyme B. Genes Cells. 1996; 1(7):673-85. DOI: 10.1046/j.1365-2443.1996.00255.x. View

16.

Jans D, Sutton V, Jans P, Froelich C, Trapani J . BCL-2 blocks perforin-induced nuclear translocation of granzymes concomitant with protection against the nuclear events of apoptosis. J Biol Chem. 1999; 274(7):3953-61. DOI: 10.1074/jbc.274.7.3953. View

17.

Bird C, Sutton V, Sun J, Hirst C, Novak A, Kumar S . Selective regulation of apoptosis: the cytotoxic lymphocyte serpin proteinase inhibitor 9 protects against granzyme B-mediated apoptosis without perturbing the Fas cell death pathway. Mol Cell Biol. 1998; 18(11):6387-98. PMC: 109224. DOI: 10.1128/MCB.18.11.6387. View

18.

Davis J, Sutton V, Smyth M, Trapani J . Dependence of granzyme B-mediated cell death on a pathway regulated by Bcl-2 or its viral homolog, BHRF1. Cell Death Differ. 2001; 7(10):973-83. DOI: 10.1038/sj.cdd.4400725. View

19.

Heibein J, Barry M, Motyka B, Bleackley R . Granzyme B-induced loss of mitochondrial inner membrane potential (Delta Psi m) and cytochrome c release are caspase independent. J Immunol. 1999; 163(9):4683-93. View

20.

Froelich C, Orth K, Turbov J, Seth P, Gottlieb R, Babior B . New paradigm for lymphocyte granule-mediated cytotoxicity. Target cells bind and internalize granzyme B, but an endosomolytic agent is necessary for cytosolic delivery and subsequent apoptosis. J Biol Chem. 1996; 271(46):29073-9. DOI: 10.1074/jbc.271.46.29073. View