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1.
Clohessy JG Zhuang J de Boer J Gil-Gómez G Brady HJ 《The Journal of biological chemistry》2006,281(9):5750-5759
Engagement of death receptors such as tumor necrosis factor-R1 and Fas brings about the cleavage of cytosolic Bid to truncated Bid (tBid), which translocates to mitochondria to activate Bax/Bak, resulting in the release of cytochrome c. The mechanism underlying the activation, however, is not fully understood. Here, we have identified the anti-apoptotic Bcl-2 family member Mcl-1 as a potent tBid-binding partner. Site-directed mutagenesis reveals that the Bcl-2 homology (BH)3 domain of tBid is essential for binding to Mcl-1, whereas all three BH domains (BH1, BH2, and BH3) of Mcl-1 are required for interaction with tBid. In vitro studies using isolated mitochondria and recombinant proteins demonstrate that Mcl-1 strongly inhibits tBid-induced cytochrome c release. In addition to its ability to interact directly with Bax and Bak, tBid also binds Mcl-1 and displaces Bak from the Mcl-1-Bak complex. Importantly, overexpression of Mcl-1 confers resistance to the induction of apoptosis by both TRAIL and tumor necrosis factor-alpha in HeLa cells, whereas targeting Mcl-1 by RNA interference sensitizes HeLa cells to TRAIL-induced apoptosis. Therefore, our study demonstrates a novel regulation of tBid by Mcl-1 through protein-protein interaction in apoptotic signaling from death receptors to mitochondria. 相似文献
2.
Head and neck squamous cell carcinoma (HNSCC) is often resistant to conventional chemotherapy and thus requires novel treatment regimens. Here, we investigated the effects of the proteasome inhibitor MG132 in combination with tumor necrosis factor-related apoptosis inducing ligand (TRAIL) or agonistic TRAIL receptor 1 (DR4)-specific monoclonal antibody, AY4, on sensitization of TRAIL- and AY4-resistant human HNSCC cell lines. Combination treatment of HNSCC cells synergistically induced apoptotic cell death accompanied by caspase-8, caspase-9, and caspase-3 activation and Bid cleavage into truncated Bid (tBid). Generation and accumulation of tBid through the cooperative action of MG132 with TRAIL or AY4 and Bik accumulation through MG132-mediated proteasome inhibition are critical to the synergistic apoptosis. In HNSCC cells, Bak was constrained by Mcl-1 and Bcl-X(L), but not by Bcl-2. Conversely, Bax did not interact with Mcl-1, Bcl-X(L), or Bcl-2. Importantly, tBid plays a major role in Bax activation, and Bik indirectly activates Bak by displacing it from Mcl-1 and Bcl-X(L), pointing to the synergistic mechanism of the combination treatment. In addition, knockdown of both Mcl-1 and Bcl-X(L) significantly sensitized HNSCC cells to TRAIL and AY4 as a single agent, suggesting that Bak constraint by Mcl-1 and Bcl-X(L) is an important resistance mechanism of TRAIL receptor-mediated apoptotic cell death. Our results provide a novel molecular mechanism for the potent synergy between MG132 proteasome inhibitor and TRAIL receptor agonists in HNSCC cells, suggesting that the combination of these agents may offer a new therapeutic strategy for HNSCC treatment. 相似文献
3.
Meng XW Lee SH Dai H Loegering D Yu C Flatten K Schneider P Dai NT Kumar SK Smith BD Karp JE Adjei AA Kaufmann SH 《The Journal of biological chemistry》2007,282(41):29831-29846
Previous studies have suggested that Mcl-1, an antiapoptotic Bcl-2 homolog that does not exhibit appreciable affinity for the caspase 8-generated C-terminal Bid fragment (tBid), diminishes sensitivity to tumor necrosis factor-alpha-related apoptosis-inducing ligand (TRAIL). This study was performed to determine the mechanism by which Mcl-1 confers TRAIL resistance and to evaluate methods for overcoming this resistance. Affinity purification/immunoblotting assays using K562 human leukemia cells, which contain Mcl-1 and Bcl-x(L) as the predominant antiapoptotic Bcl-2 homologs, demonstrated that TRAIL treatment resulted in binding of tBid to Bcl-x(L) but not Mcl-1. In contrast, TRAIL caused increased binding between Mcl-1 and Bak that was diminished by treatment with the caspase 8 inhibitor N-(N(alpha)-acetylisoleucylglutamylthreonyl) aspartic acid (O-methyl ester)-fluoromethyl ketone (IETD(OMe)-fmk) or the c-Jun N-terminal kinase inhibitor SP600125. In addition, TRAIL caused increased binding of Bim and Puma to Mcl-1 that was inhibited by IETD(OMe)-fmk but not SP600125. Further experiments demonstrated that down-regulation of Mcl-1 by short hairpin RNA or the kinase inhibitor sorafenib increased TRAIL-induced Bak activation and death ligand-induced apoptosis in a wide variety of neoplastic cell lines as well as clinical acute myelogenous leukemia specimens. Collectively, these observations not only suggest a model in which Mcl-1 confers TRAIL resistance by serving as a buffer for Bak, Bim, and Puma, but also identify sorafenib as a potential modulator of TRAIL sensitivity. 相似文献
4.
The mitogen-activated protein kinase pathway can inhibit TRAIL-induced apoptosis by prohibiting association of truncated Bid with mitochondria 总被引:2,自引:0,他引:2
Ortiz-Ferrón G Tait SW Robledo G de Vries E Borst J López-Rivas A 《Cell death and differentiation》2006,13(11):1857-1865
Breast cancer cells often show increased activity of the mitogen-activated protein kinase (MAPK) pathway. We report here that this pathway reduces their sensitivity to death ligand tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and present the underlying mechanism. Activation of protein kinase C (PKC) inhibited TRAIL-induced apoptosis in a protein synthesis-independent manner. Deliberate activation of MAPK was also inhibitory. In digitonin-permeabilized cells, PKC activation interfered with the capacity of recombinant truncated (t)Bid to release cytochrome c from mitochondria. MAPK activation did not affect TRAIL or tumor necrosis factor (TNF)alpha-induced Bid cleavage. However, it did inhibit translocation of (t)Bid to mitochondria as determined both by subcellular fractionation analysis and confocal microscopy. Steady state tBid mitochondrial localization was prohibited by activation of the MAPK pathway, also when the Bcl-2 homology domain 3 (BH3) domain of tBid was disrupted. We conclude that the MAPK pathway inhibits TRAIL-induced apoptosis in MCF-7 cells by prohibiting anchoring of tBid to the mitochondrial membrane. This anchoring is independent of its interaction with resident Bcl-2 family members. 相似文献
5.
In many mammalian cell types, engagement of the TRAIL/Apo2L death receptors DR4 and DR5 alters mitochondrial physiology, thereby promoting the release of pro-apoptotic proteins normally contained within this organelle. A contemporary view of this process is that in so-called type II cells death receptor-activated caspase-8 cleaves the Bcl-2 family member Bid, which generates a truncated Bid fragment that collaborates with Bax, another Bcl-2 relative, to promote the release of mitochondrial factors necessary for activation of executioner caspases and apoptosis. Here we show that in some type II cells caspase-2 is necessary for optimal TRAIL-mediated cleavage of Bid. Down-regulation of caspase-2 using RNA interference significantly inhibited TRAIL-induced apoptosis. Analysis of the TRAIL proteolytic cascade following gene silencing of specific pathway components revealed that caspase-2 is necessary for efficient cleavage of Bid; however, caspase-2 proteolytic processing, which occurs downstream of Bax, is not necessary for its role in Bid cleavage. 相似文献
6.
Falko Lange Bettina Brandt Markus Tiedge Ludwig Jonas Udo Jeschke Ralf Pöhland Hermann Walzel 《Histochemistry and cell biology》2009,132(2):211-223
Galectin-1 (gal-1) triggers T cell death by several distinct intracellular pathways including the activation of the death-receptor
pathway. The aim of this study was to investigate whether gal-1 induced activation of the death-receptor pathway in Jurkat
T lymphocytes mediates apoptosis via the mitochondrial pathway linked by truncated Bid (tBid). We demonstrate that gal-1 induced
proteolytic cleavage of the death agonist Bid, a member of the Bcl-2/Bcl-xL family and a substrate of activated caspase-8,
was inhibited by caspase-8 inhibitor II (Z-IETD-FMK). Downstream of Bid, gal-1 stimulated mitochondrial cytochrome c release as well as the activation and proteolytic processing of initiator procaspase-9 were effectively decreased by caspase-8
inhibitor II. Blocking of gal-1 induced cleavage of effector procaspase-3 by caspase-8 inhibitor II as well as by caspase-9
inhibitors I (Z-LEHD-FMK) and III (Ac-LEHD-CMK) indicates that receptor and mitochondrial pathways converged in procaspase-3
activation and contribute to proteolytic processing of effector procaspase-6 and -7. Western blot analyses and immunofluorescence
staining revealed that exposure of Jurkat T cells to gal-1 resulted in the cleavage of the DNA-repair enzyme poly (ADP-ribose)
polymerase, cytoskeletal α-fodrin, and nuclear lamin A as substrates of activated caspases. Our data demonstrate that Bid
provides a connection between the death receptor and the mitochondrial pathway of gal-1 induced apoptosis in human Jurkat
T lymphocytes. 相似文献
7.
Kawanishi M Tada-Oikawa S Kawanishi S 《Biochemical and biophysical research communications》2002,297(3):682-687
We have previously reported that TNF-related apoptosis inducing ligand (TRAIL) causes cleavage of Bid via activation of caspase-8 and the loss of mitochondrial membrane potential (DeltaPsim), resulting in apoptosis. Experiments with BJAB clones expressing Epstein-Barr virus (EBV) anti-apoptotic protein BHRF1 showed that BHRF1 drastically inhibited TRAIL-mediated apoptosis. Although Western blot analysis demonstrated that TRAIL-induced Bid cleavage was not inhibited by BHRF1, the decrease in DeltaPsim caused by TRAIL was effectively blocked by BHRF1. These findings suggest that in BJAB cells, BHRF1 acts downstream of Bid cleavage and upstream of mitochondrial damage, resulting in inhibition of TRAIL-induced apoptosis. 相似文献
8.
Exposure of cells to hyperthermia is known to induce apoptosis, although the underlying mechanisms are only partially understood. Here, we examine the molecular requirements necessary for heat-induced apoptosis using genetically modified Jurkat T-lymphocytes. Cells stably overexpressing Bcl-2/Bcl-x(L) or stably depleted of Apaf-1 were completely resistant to heat-induced apoptosis, implicating the involvement of the mitochondria-mediated pathway. Pretreatment of wild-type cells with the cell-permeable biotinylated general caspase inhibitor b-VAD-fmk (biotin-Val-Ala-Asp(OMe)-CH(2)F) both inhibited heat-induced apoptosis and affinity-labeled activated initiator caspase-2, -8, and -9. Despite this finding, however, cells engineered to be deficient in caspase-8, caspase-2, or the caspase-2 adaptor protein RAIDD (receptor-interacting protein (RIP)-associated Ich-1/CED homologous protein with death domain) remained susceptible to heat-induced apoptosis. Additionally, b-VAD-fmk failed to label any activated initiator caspase in Apaf-1-deficient cells exposed to hyperthermia. Cells lacking Apaf-1 or the pro-apoptotic BH3-only protein Bid exhibited lower levels of heat-induced Bak activation, cytochrome c release, and loss of mitochondrial membrane potential, although cleavage of Bid to truncated Bid (tBid) occurred downstream of caspase-9 activation. Combined, the data suggest that caspase-9 is the critical initiator caspase activated during heat-induced apoptosis and that tBid may function to promote cytochrome c release during this process as part of a feed-forward amplification loop. 相似文献
9.
Mary E. Shawgo Shary N. Shelton John D. Robertson 《The Journal of biological chemistry》2009,284(48):33447-33455
Activation of executioner caspases during receptor-mediated apoptosis in type II cells requires the engagement of the mitochondrial apoptotic pathway. Although it is well established that recruitment of mitochondria in this context involves the cleavage of Bid to truncated Bid (tBid), the precise post-mitochondrial signaling responsible for executioner caspase activation is controversial. Here, we used distinct clones of type II Jurkat T-lymphocytes in which the mitochondrial apoptotic pathway had been inhibited to investigate the molecular requirements necessary for Fas-induced apoptosis. Cells overexpressing either Bcl-2 or Bcl-xL were protected from apoptosis induced by agonistic anti-Fas antibody. By comparison, Apaf-1-deficient Jurkat cells were sensitive to anti-Fas, exhibiting Bid cleavage, Bak activation, the release of cytochrome c and Smac, and activation of executioner caspase-3. Inhibiting downstream caspase activation with the pharmacological inhibitor Z-DEVD-fmk or by expressing the BIR1/BIR2 domains of X-linked inhibitor of apoptosis protein (XIAP) decreased all anti-Fas-induced apoptotic changes. Additionally, pretreatment of Bcl-xL-overexpressing cells with a Smac mimetic sensitized these cells to Fas-induced apoptosis. Combined, our findings strongly suggest that Fas-mediated activation of executioner caspases and induction of apoptosis do not depend on apoptosome-mediated caspase-9 activation in prototypical type II cells. 相似文献
10.
Bcl-2 family member Bfl-1/A1 sequesters truncated bid to inhibit is collaboration with pro-apoptotic Bak or Bax 总被引:11,自引:0,他引:11
Werner AB de Vries E Tait SW Bontjer I Borst J 《The Journal of biological chemistry》2002,277(25):22781-22788
Following caspase-8 mediated cleavage, a carboxyl-terminal fragment of the BH3 domain-only Bcl-2 family member Bid transmits the apoptotic signal from death receptors to mitochondria. In a screen for possible regulators of Bid, we defined Bfl-1/A1 as a potent Bid interacting protein. Bfl-1 is an anti-apoptotic Bcl-2 family member, whose preferential expression in hematopoietic cells and endothelium is controlled by inflammatory stimuli. Its mechanism of action is unknown. We find that Bfl-1 associates with both full-length Bid and truncated (t)Bid, via the Bid BH3 domain. Cellular expression of Bfl-1 confers protection against CD95- and Trail receptor-induced cytochrome c release. In vitro assays, using purified mitochondria and recombinant proteins, demonstrate that Bfl-1 binds full-length Bid, but does not interfere with its processing by caspase-8, or with its mitochondrial association. Confocal microscopy supports that Bfl-1, which at least in part constitutively localizes to mitochondria, does not impede tBid translocation. However, Bfl-1 remains tightly and selectively bound to tBid and blocks collaboration between tBid and Bax or Bak in the plane of the mitochondrial membrane, thereby preventing mitochondrial apoptotic activation. Lack of demonstrable interaction between Bfl-1 and Bak or Bax in the mitochondrial membrane suggests that Bfl-1 generally prevents the formation of a pro-apoptotic complex by sequestering BH3 domain-only proteins. 相似文献
11.
Identification of mechanisms of modulation of the TNF-related apoptosis-inducing ligand (TRAIL)-induced apoptosis is important for its potential use in anticancer therapy. Ethanol can induce cell death in vitro and in vivo by different signalling pathways. Its effect in combination with death ligands is unknown. We investigated how ethanol modulates the effects of TRAIL in colon cancer cells. After combined TRAIL and ethanol treatment, a potentiation of caspase-8, -9, -3 activation, a proapoptotic Bid protein cleavage, a decrease of mitochondrial membrane potential, a complete poly(ADP)ribose polymerase cleavage, and disappearance of antiapoptotic Mcl-1 protein were demonstrated. Ethanol acts as a potent agent sensitizing colon cancer cells to TRAIL-induced apoptosis. 相似文献
12.
Previously we have shown that interferon (IFN)-α induced apoptosis is predominantly mediated by the upregulation of tumor necrosis factor related apoptosis-inducing ligand (TRAIL) via the caspase-8 pathway. It was also shown that recruitment of mitochondria in IFN-α induced apoptosis involves the cleavage of BH3 interacting domain death agonist (Bid) to truncated Bid (tBid). In the present study, we demonstrate that tBid induced by IFN-α2a activates mitochondrial Bak to trigger the loss of mitochondrial membrane integrity, consequently causing release of apoptosis-inducing factor (AIF) in ovarian cancer cells, OVCAR3. AIF translocates from the mitochondria to the nucleus and induces nuclear fragmentation and cell death. Both a small molecule Bid inhibitor (BI-6C9) or Bid-RNA interference (RNAi) preserved mitochondrial membrane potential, prevented nuclear translocation of AIF, and abrogated IFN-α2a-induced cell death. Cell death induced by tBid was inhibited by AIF-RNAi, indicating that caspase-independent AIF signaling is the main pathway through which Bid mediates cell death. This was further supported by experiments showing that BI-6C9 did not prevent the release of cytochrome c from mitochondria to cytosol, while the release of AIF was prevented. In conclusion, IFN-α2a-induced apoptosis is mediated via the mitochondria-associated pathway involving the cleavage of Bid followed by AIF release that involves Bak activation and translocation of AIF from the mitochondria to the nucleus in OVCAR3 cells. 相似文献
13.
Bernhard Gillissen Jana Wendt Antje Richter Anja Richter Annika Müer Tim Overkamp Nina Gebhardt Robert Preissner Claus Belka Bernd D?rken Peter T. Daniel 《The Journal of cell biology》2010,188(6):851-862
Tumor necrosis factor (α)–related apoptosis-inducing ligand (TRAIL) is a promising anticancer agent that preferentially kills tumor cells with limited cytotoxicity to nonmalignant cells. However, signaling from death receptors requires amplification via the mitochondrial apoptosis pathway (type II) in the majority of tumor cells. Thus, TRAIL-induced cell death entirely depends on the proapoptotic Bcl-2 family member Bax, which is often lost as a result of epigenetic inactivation or mutations. Consequently, Bax deficiency confers resistance against TRAIL-induced apoptosis. Despite expression of Bak, Bax-deficient cells are resistant to TRAIL-induced apoptosis. In this study, we show that the Bax dependency of TRAIL-induced apoptosis is determined by Mcl-1 but not Bcl-xL. Both are antiapoptotic Bcl-2 family proteins that keep Bak in check. Nevertheless, knockdown of Mcl-1 but not Bcl-xL overcame resistance to TRAIL, CD95/FasL and tumor necrosis factor (α) death receptor ligation in Bax-deficient cells, and enabled TRAIL to activate Bak, indicating that Mcl-1 rather than Bcl-xL is a major target for sensitization of Bax-deficient tumors for death receptor–induced apoptosis via the Bak pathway. 相似文献
14.
A caspase-8-independent component in TRAIL/Apo-2L-induced cell death in human rhabdomyosarcoma cells 总被引:6,自引:0,他引:6
Petak I Vernes R Szucs KS Anozie M Izeradjene K Douglas L Tillman DM Phillips DC Houghton JA 《Cell death and differentiation》2003,10(6):729-739
Tumor necrosis factor related apoptosis inducing ligand (TRAIL) belongs to the Tumor necrosis factor (TNF) family of death-inducing ligands, and signaling downstream of TRAIL ligation to its receptor(s) remains to be fully elucidated. Components of the death-inducing signaling complex (DISC) and TRAIL signaling downstream of receptor activation were examined in TRAIL - sensitive and -resistant models of human rhabdomyosarcoma (RMS). TRAIL ligation induced DISC formation in TRAIL-sensitive (RD, Rh18, Rh30) and TRAIL-resistant RMS (Rh28, Rh36, Rh41), with recruitment of FADD and procaspase-8. In RD cells, overexpression of dominant-negative FADD (DNFADD) completely abolished TRAIL-induced cell death in contrast to dominant-negative caspase- 8 (DNC8), which only partially inhibited TRAIL-induced apoptosis, growth inhibition, or loss in clonogenic survival. DNC8 did not inhibit the cleavage of Bid or the activation of Bax. Overexpression of Bcl-2 or Bcl-xL inhibited TRAIL-induced apoptosis, growth inhibition, and loss in clonogenic survival. Bcl-2 and Bcl-xL, but not DNC8, inhibited TRAIL-induced Bax activation. Bcl-xL did not inhibit the early activation of caspase-8 (<4 h) but inhibited cleavage of Bid, suggesting that Bid is cleaved downstream of the mitochondria, independent of caspase-8. Exogenous addition of sphingosine also induced activation of Bax via a caspase-8-and Bid-independent mechanism. Further, inhibition of sphingosine kinase completely protected cells from TRAIL-induced apoptosis. Data demonstrate that in RMS cells, the TRAIL signaling pathway circumvents caspase-8 activation of Bid upstream of the mitochondria and that TRAIL acts at the level of the mitochondria via a mechanism that may involve components of the sphingomyelin cycle. 相似文献
15.
TRAIL causes cleavage of bid by caspase-8 and loss of mitochondrial membrane potential resulting in apoptosis in BJAB cells. 总被引:13,自引:0,他引:13
H Yamada S Tada-Oikawa A Uchida S Kawanishi 《Biochemical and biophysical research communications》1999,265(1):130-133
A new member of the TNF family, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), has been shown to induce apoptosis. However, the mechanism for TRAIL-induced apoptosis remains to be clarified. SDS-PAGE and Western blot analysis showed that cleavage of Bid was induced by a 1-h incubation of BJAB cells with TRAIL and was blocked by a caspase-8 inhibitor. Flow cytometry demonstrated that loss of mitochondrial membrane potential in BJAB cells began about 1.5 h after the treatment with TRAIL and was apparent at 2 h in comparison with the control. DNA ladder formation, which is characteristic for apoptosis, in the cells treated with TRAIL was detected at 2 h and observed most effectively at 3 h. The time course study suggests that TRAIL causes cleavage of Bid via activation of caspase-8, subsequently the loss of mitochondrial membrane potential, resulting in apoptosis in BJAB cells. 相似文献
16.
Shary N. Shelton Mary E. Shawgo John D. Robertson 《The Journal of biological chemistry》2009,284(17):11247-11255
The extent to which the BH3-only protein Bid is important for intrinsic
(mitochondria-mediated) apoptotic cell death induced by genotoxic stress
remains controversial. In the present study, we examine this issue using a
panel of gene-manipulated Bax-deficient Jurkat T-lymphocytes. Cells stably
depleted of Bid were far less sensitive than control-transfected cells to
etoposide-induced apoptosis. In particular, drug-induced Bak activation,
cytochrome c release, loss of mitochondrial membrane potential, and
caspase activation were all decreased in cells lacking Bid. Reconstitution
experiments using recombinant proteins and permeabilized Bid-deficient cells
demonstrated that truncated Bid (tBid), but not full-length Bid, potently
induced Bak activation and the release of cytochrome c. Further,
caspase-8-deficient Jurkat cells efficiently cleaved Bid and were sensitive to
drug-induced apoptosis. By comparison, Apaf-1-deficient cells, as well as
cells overexpressing full-length X-linked inhibitor of apoptosis protein
(XIAP) or the BIR1/BIR2 domains of XIAP, failed to cleave Bid in response to
genotoxic stress. These data suggest that tBid plays an important regulatory
role in the execution of DNA damage-induced cytochrome c release and
apoptosis. However, the fact that cleavage of Bid to tBid is mediated by
executioner caspases suggests that a self-amplifying feed forward loop
involving caspases, Bid, and mitochondria may help determine irreversible
commitment to apoptosis.Apoptosis is an active form of cell death that plays an essential role
during normal embryonic development and in the maintenance of tissue
homeostasis in the adult organism
(1). Consequently,
dysregulation of apoptosis has been implicated as a contributing factor to the
onset of different pathological conditions, including cancer. In addition, it
is now generally accepted that many genotoxic anticancer drugs are effective
against tumor cells for their ability to induce mitochondria-mediated
apoptosis (2). Similarly,
mutations or the altered expression of pro- and anti-apoptotic proteins can
contribute to the development of drug resistance.Execution of apoptosis is mediated by a family of cysteine-dependent
aspartate-specific proteases (caspases). During true mitochondria-mediated
apoptosis, members of the Bcl-2 family of proteins are the primary regulators
of caspase activation for their role in controlling mitochondrial outer
membrane permeabilization
(MOMP)2
(3). The process of MOMP
results in the release of cytochrome c, second mitochondria-derived
activator of caspase (Smac, also known as DIABLO), and Omi (also known as
HtrA2) into the cytosol where they converge to promote the activation of
caspase-9 within the apoptotic protease-activating factor-1 (Apaf-1)
apoptosome complex. The Bcl-2 family contains proteins with opposing
functions, and it is generally thought that the induction of MOMP requires the
activation of either Bak or Bax triggered by a Bcl-2 homology 3 (BH3)-only
protein
(4–6).
Indeed, evidence in the literature indicates that cells lacking either Bak or
Bax exhibit only subtle defects in MOMP, whereas doubly deficient cells are
often found to be highly resistant to mitochondria-mediated apoptosis
(7,
8).At present, there are two models for the activation of Bax or Bak by
BH3-only proteins. One model argues that BH3-only proteins function as either
“sensitizer” (e.g. Bad and Noxa) or
“activator” proteins (e.g. truncated Bid (tBid), Bim, and
perhaps Puma) (9). In this
scenario, a sensitizer protein is needed to displace an activator protein from
a prosurvival protein (e.g. Bcl-2, Bcl-xL, or Mcl-1) to
activate Bak or Bax. The second model argues that BH3-only proteins bind and
inhibit the function of prosurvival Bcl-2 proteins, which normally bind to and
inhibit Bak and Bax (10,
11). Of the seven or so known
BH3-only proteins (6), Bid is
unique in that it requires post-translational modification for activation,
most notably involving caspase-8-mediated cleavage to tBid
(12–14).
Bid normally resides in the cytosol and possibly the nucleus
(15). Upon being cleaved, the
C-terminal fragment (tBid) is myristoylated at its newly exposed N terminus,
translocates to the outer mitochondrial membrane (OMM), and/or activates Bak
or Bax protein (16). Recently,
it was shown that the N-terminal cleavage fragment of Bid is quickly
ubiquitinated for degradation and that this degradation is necessary for the
pro-apoptotic function of tBid
(17). The same study also
concluded that, although full-length Bid is capable of translocating to the
OMM, it is not able to induce MOMP on its own
(17). A well characterized
example of tBid involvement during apoptosis is in the engagement of the
mitochondrial apoptotic pathway in so-called type II cells upon activation of
the extrinsic pathway
(18).Here, we have investigated whether Bid plays a functional role in the
induction of MOMP during apoptosis in response to the genotoxic anticancer
drug etoposide. To that end, we used Bax-deficient Jurkat cells that are
stably depleted of Bid and evaluated the extent to which these cells underwent
drug-induced MOMP. In addition, Jurkat clones in which the intrinsic pathway
had been inhibited due to the stable knockdown of Apaf-1 or the overexpression
of full-length XIAP or the baculoviral IAP repeats 1 and 2 (BIR1/BIR2) of XIAP
were used to gain insight into the molecular requirements necessary for
cleavage of Bid to tBid during drug-induced apoptosis. Strikingly, the data
showed that etoposide-induced apoptosis was decreased in Bid-deficient Jurkat
cells. In particular, cells lacking Bid expression exhibited decreased Bak
activation, cytochrome c release, loss of mitochondrial membrane
potential (ΔΨ), and caspase activation. Further, incubation of
permeabilized Bid-deficient cells with recombinant tBid, but not full-length
Bid, induced Bak dimerization and cytochrome c release.
Significantly, we also found that cleavage of Bid to tBid occurred strictly
downstream of Apaf-1 by a mechanism that required active executioner
caspases. 相似文献
17.
Inhibition of Bid-induced apoptosis by Bcl-2. tBid insertion,Bax translocation,and Bax/Bak oligomerization suppressed 总被引:11,自引:0,他引:11
Bcl-2 family proteins are important regulators of apoptosis. They can be pro-apoptotic (e.g. Bid, Bax, and Bak) or anti-apoptotic (e.g. Bcl-2 and Bcl-x(L)). The current study examined Bid-induced apoptosis and its inhibition by Bcl-2. Transfection of Bid led to apoptosis in HeLa cells. In these cells, Bid was processed into active forms of truncated Bid or tBid. Following processing, tBid translocated to the membrane-bound organellar fraction. Bcl-2 co-transfection inhibited Bid-induced apoptosis but did not prevent Bid processing or tBid translocation. On the other hand, Bcl-2 blocked the release of mitochondrial cytochrome c in Bid-transfected cells, suggesting actions at the mitochondrial level. Alkaline treatment stripped off tBid from the membrane-bound organellar fraction of Bid plus Bcl-2-co-transfected cells, but not from cells transfected with only Bid, suggesting inhibition of tBid insertion into mitochondrial membranes by Bcl-2. Bcl-2 also prevented Bid-induced Bax translocation from cytosol to the membrane-bound organellar fraction. Finally, Bcl-2 diminished Bid-induced oligomerization of Bax and Bak within the membrane-bound organellar fraction, shown by cross-linking experiments. In conclusion, Bcl-2 inhibited Bid-induced apoptosis at the mitochondrial level by blocking cytochrome c release, without suppressing Bid processing or activation. Critical steps blocked by Bcl-2 included tBid insertion, Bax translocation, and Bax/Bak oligomerization in the mitochondrial membranes. 相似文献
18.
Fran?ois Gonzalvez Fabrizio Pariselli Olivier Jalmar Pauline Dupaigne Franck Sureau Marc Dellinger Eric A. Hendrickson Sophie Bernard Patrice X. Petit 《PloS one》2010,5(2)
Background
The pro-apoptotic effector Bid induces mitochondrial apoptosis in synergy with Bax and Bak. In response to death receptors activation, Bid is cleaved by caspase-8 into its active form, tBid (truncated Bid), which then translocates to the mitochondria to trigger cytochrome c release and subsequent apoptosis. Accumulating evidence now indicate that the binding of tBid initiates an ordered sequences of events that prime mitochondria from the action of Bax and Bak: (1) tBid interacts with mitochondria via a specific binding to cardiolipin (CL) and immediately disturbs mitochondrial structure and function idependently of its BH3 domain; (2) Then, tBid activates through its BH3 domain Bax and/or Bak and induces their subsequent oligomerization in mitochondrial membranes. To date, the underlying mechanism responsible for targeting tBid to mitochondria and disrupting mitochondrial bioenergetics has yet be elucidated.Principal Findings
The present study investigates the mechanism by which tBid interacts with mitochondria issued from mouse hepatocytes and perturbs mitochondrial function. We show here that the helix αH6 is responsible for targeting tBid to mitochondrial CL and disrupting mitochondrial bioenergetics. In particular, αH6 interacts with mitochondria through electrostatic interactions involving the lysines 157 and 158 and induces an inhibition of state-3 respiration and an uncoupling of state-4 respiration. These changes may represent a key event that primes mitochondria for the action of Bax and Bak. In addition, we also demonstrate that tBid required its helix αH6 to efficiently induce cytochrome c release and apoptosis.Conclusions
Our findings provide new insights into the mechanism of action of tBid, and particularly emphasize the importance of the interaction of the helix αH6 with CL for both mitochondrial targeting and pro-apoptotic activity of tBid. These support the notion that tBid acts as a bifunctional molecule: first, it binds to mitochondrial CL via its helix αH6 and destabilizes mitochondrial structure and function, and then it promotes through its BH3 domain the activation and oligomerization of Bax and/or Bak, leading to cytochrome c release and execution of apoptosis. Our findings also imply an active role of the membrane in modulating the interactions between Bcl-2 proteins that has so far been underestimated. 相似文献19.
Söderström TS Poukkula M Holmström TH Heiskanen KM Eriksson JE 《Journal of immunology (Baltimore, Md. : 1950)》2002,169(6):2851-2860
Fas ligand and TNF-related apoptosis-inducing ligand (TRAIL) induce apoptosis in many different cell types. Jurkat T cells die rapidly by apoptosis after treatment with either ligand. We have previously shown that mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) can act as a negative regulator of apoptosis mediated by the Fas receptor. In this study we examined whether MAPK/ERK can also act as a negative regulator of apoptosis induced by TRAIL. Activated Jurkat T cells were efficiently protected from TRAIL-induced apoptosis. The protection was shown to be MAPK/ERK dependent and independent of protein synthesis. MAPK/ERK suppressed TRAIL-induced apoptosis upstream of the mitochondrial amplification loop because mitochondrial depolarization and release of cytochrome c were inhibited. Furthermore, caspase-8-mediated relocalization and activation of Bid, a proapoptotic member of the Bcl family, was also inhibited by the MAPK/ERK signaling. The protection occurred at the level of the apoptotic initiator caspase-8, as the cleavage of caspase-8 was inhibited but the assembly of the death-inducing signaling complex was unaffected. Both TRAIL and Fas ligand have been suggested to regulate the clonal size and persistence of different T cell populations. Our previous results indicate that MAPK/ERK protects recently activated T cells from Fas receptor-mediated apoptosis during the initial phase of an immune response before the activation-induced cell death takes place. The results of this study show clearly that MAPK/ERK also participates in the inhibition of TRAIL-induced apoptosis after T cell activation. 相似文献
20.
Soo-Jung Park Mi-Ju Kim Hee-Young Sohn Chi-Dug Kang Sun-Hee Kim 《Experimental cell research》2009,315(11):1809-77
TNF-related apoptosis-inducing ligand (TRAIL) is a pro-apoptotic cytokine that is capable of inducing apoptosis in a wide variety of cancer cells but not in normal cells. Although many cancer cells are sensitive to TRAIL-induced apoptosis, chronic myeloid leukemia (CML) develops resistance to TRAIL. In this study, we investigated whether apicidin, a novel histone deacetylase inhibitor, could overcome the TRAIL resistance in CML-derived K562 cells. Compared to treatment with apicidin or TRAIL alone, cotreatment with apicidin and TRAIL-induced apoptosis synergistically in K562 cells. This combination led to activation of caspase-8 and Bcl-2 interacting domain (Bid), resulting in the cytosolic accumulation of cytochrome c from mitochondria as well as an activation of caspase-3. Treatment with apicidin resulted in down-regulation of Bcr-Abl and inhibition of its downstream target, PI3K/AKT-NF-κB pathway. In addition, apicidin decreased the level of NF-κB-dependent Bcl-xL, leading to caspase activation and Bid cleavage. These results suggest that apicidin may sensitize K562 cells to TRAIL-induced apoptosis through caspase-dependent mitochondrial pathway by regulating expression of Bcr-Abl and its related anti-apoptotic proteins. Therefore, the present study suggests that combination of apicidin and TRAIL may be an effective strategy for treating TRAIL-resistant Bcr-Abl expressing CML cells. 相似文献