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The EMBO Journal Feb 2007Mitochondrial outer membrane permeabilization (MOMP) and release of mitochondrial intermembrane proteins like cytochrome c are critical steps in the control of... (Comparative Study)
Comparative Study
Mitochondrial outer membrane permeabilization (MOMP) and release of mitochondrial intermembrane proteins like cytochrome c are critical steps in the control of apoptosis. Previous work has shown that MOMP depends on the functionally redundant multidomain proapoptotic proteins, Bak and Bax. Here we demonstrate that Bak and Bax are functionally non-redundant during Neisseria gonorrhoeae (Ngo)- and cisplatin-induced apoptosis. While the activation of Bak is caspase independent Bax activation needs Bak and active caspases. Silencing of either Bak or Bax resists both Ngo- and cisplatin- but not TNFalpha-induced apoptosis. Activation of Bak is required to release cytochrome c from the mitochondria; however, Bax is still required to activate effector caspases. Thus, both Bak and Bax are necessary to accomplish DNA damage and Ngo-induced apoptosis.
Topics: Apoptosis; Blotting, Western; Caspases; Cell Fractionation; DNA Damage; DNA Primers; Flow Cytometry; HeLa Cells; Humans; Immunoprecipitation; Intracellular Membranes; Microscopy, Fluorescence; Mitochondria; Neisseria gonorrhoeae; Permeability; RNA Interference; bcl-2 Homologous Antagonist-Killer Protein; bcl-2-Associated X Protein
PubMed: 17235284
DOI: 10.1038/sj.emboj.7601533 -
Cell Death and Differentiation Oct 2022Apoptosis acts in defense against microbial infection, and many infectious agents have developed strategies to inhibit host cell apoptosis. The human pathogen Chlamydia...
Apoptosis acts in defense against microbial infection, and many infectious agents have developed strategies to inhibit host cell apoptosis. The human pathogen Chlamydia trachomatis (Ctr) is an obligate intracellular bacterium that strongly inhibits mitochondrial apoptosis of its human host cell but there is no agreement how the bacteria achieve this. We here provide a molecular analysis of chlamydial apoptosis-inhibition in infected human cells and demonstrate that the block of apoptosis occurs during the activation of the effectors of mitochondrial apoptosis, Bak and Bax. We use small-molecule Bcl-2-family inhibitors and gene targeting to show that previous models cannot explain the anti-apoptotic effect of chlamydial infection. Although the anti-apoptotic Bcl-2-family protein Mcl-1 was strongly upregulated upon infection, Mcl-1-deficient cells and cells where Mcl-1 was pharmacologically inactivated were still protected. Ctr-infection could inhibit both Bax- and Bak-induced apoptosis. Apoptotic Bax-oligomerization and association with the outer mitochondrial membrane was reduced upon chlamydial infection. Infection further inhibited apoptosis induced conformational changes of Bak, as evidenced by changes to protease sensitivity, oligomerization and release from the mitochondrial porin VDAC2. Mitochondria isolated from Ctr-infected cells were protected against the pro-apoptotic Bcl-2-family proteins Bim and tBid but this protection was lost upon protease digestion. However, the protective effect of Ctr-infection was reduced in cells lacking the Bax/Bak-regulator VDAC2. We further found that OmpA, a porin of the outer membrane of Ctr, associated upon experimental expression with mitochondria and inhibited apoptosis, phenocopying the effect of the infection. These results identify a novel way of apoptosis inhibition, involving only the most downstream modulator of mitochondrial apoptosis and suggest that Chlamydia has a protein dedicated to the inhibition of apoptosis to secure its survival in human cells.
Topics: Apoptosis; Apoptosis Regulatory Proteins; Chlamydia trachomatis; Humans; Myeloid Cell Leukemia Sequence 1 Protein; Peptide Hydrolases; Proto-Oncogene Proteins c-bcl-2; bcl-2 Homologous Antagonist-Killer Protein; bcl-2-Associated X Protein
PubMed: 35397654
DOI: 10.1038/s41418-022-00995-0 -
Cell Death and Differentiation Jan 2018The BCL-2 family of proteins controls cell death primarily by direct binding interactions that regulate mitochondrial outer membrane permeabilization (MOMP) leading to... (Review)
Review
The BCL-2 family of proteins controls cell death primarily by direct binding interactions that regulate mitochondrial outer membrane permeabilization (MOMP) leading to the irreversible release of intermembrane space proteins, subsequent caspase activation and apoptosis. The affinities and relative abundance of the BCL-2 family proteins dictate the predominate interactions between anti-apoptotic and pro-apoptotic BCL-2 family proteins that regulate MOMP. We highlight the core mechanisms of BCL-2 family regulation of MOMP with an emphasis on how the interactions between the BCL-2 family proteins govern cell fate. We address the critical importance of both the concentration and affinities of BCL-2 family proteins and show how differences in either can greatly change the outcome. Further, we explain the importance of using full-length BCL-2 family proteins (versus truncated versions or peptides) to parse out the core mechanisms of MOMP regulation by the BCL-2 family. Finally, we discuss how post-translational modifications and differing intracellular localizations alter the mechanisms of apoptosis regulation by BCL-2 family proteins. Successful therapeutic intervention of MOMP regulation in human disease requires an understanding of the factors that mediate the major binding interactions between BCL-2 family proteins in cells.
Topics: Apoptosis; Apoptosis Regulatory Proteins; Cell Membrane Permeability; Mitochondrial Membranes; Protein Processing, Post-Translational; Proto-Oncogene Proteins c-bcl-2; bcl-2 Homologous Antagonist-Killer Protein; bcl-2-Associated X Protein
PubMed: 29149100
DOI: 10.1038/cdd.2017.186 -
Cell Death & Disease Jun 2013The multi-BCL-2 homology domain pro-apoptotic BCL-2 family members BAK and BAX have critical roles in apoptosis. They are essential for mitochondrial outer-membrane...
The multi-BCL-2 homology domain pro-apoptotic BCL-2 family members BAK and BAX have critical roles in apoptosis. They are essential for mitochondrial outer-membrane permeabilization, leading to the release of apoptogenic factors such as cytochrome-c, which promote activation of the caspase cascade and cellular demolition. The BOK protein has extensive amino-acid sequence similarity to BAK and BAX and is expressed in diverse cell types, particularly those of the female reproductive tissues. The BOK-deficient mice have no readily discernible abnormalities, and its function therefore remains unresolved. We hypothesized that BOK may exert functions that overlap with those of BAK and/or BAX and examined this by generating Bok(-/-)Bak(-/-) and Bok(-/-)Bax(-/-) mice. Combined loss of BOK and BAK did not elicit any noticeable defects, although it remains possible that BOK and BAK have critical roles in developmental cell death that overlap with those of BAX. In most tissues examined, loss of BOK did not exacerbate the abnormalities caused by loss of BAX, such as defects in spermatogenesis or the increase in neuronal populations in the brain and retina. Notably, however, old Bok(-/-)Bax(-/-) females had abnormally increased numbers of oocytes from different stages of development, indicating that BOK may have a pro-apoptotic function overlapping with that of BAX in age-related follicular atresia.
Topics: Animals; Apoptosis; Brain; Cell Survival; Cells, Cultured; Female; Lymphocytes; Male; Mice; Mice, 129 Strain; Mice, Inbred C57BL; Mice, Knockout; Myeloid Cells; Ovarian Follicle; Proto-Oncogene Proteins c-bcl-2; Retina; Spleen; Testis; bcl-2 Homologous Antagonist-Killer Protein; bcl-2-Associated X Protein
PubMed: 23744350
DOI: 10.1038/cddis.2013.176 -
Anticancer Research Oct 2009EPO (erythropoietin) counteracts p53-dependent apoptosis. EPO, which acts via its receptor EPOR, protects cells and inhibits apoptosis in normal cells and some cancer...
BACKGROUND
EPO (erythropoietin) counteracts p53-dependent apoptosis. EPO, which acts via its receptor EPOR, protects cells and inhibits apoptosis in normal cells and some cancer tissues by severe down-regulation of Bak. We aimed to investigate the relationship between p53 and Bak expression and EPO and EPOR in human colorectal carcinomas.
MATERIALS AND METHODS
The expression of p53 was compared with Bak, EPO and EPOR in 96 colorectal carcinomas by means of immunohistochemistry.
RESULTS
Purely nuclear p53 was significantly higher expressed in the moderately differentiated cancers in comparison with the poorly differentiated ones (p=0.007). P53 expression did not correlate with cytoplasmic markers: Bak, EPO and EPOR, but EPO and EPOR were significantly associated with Bak expression (p<0.001, r=0.524 and p<0.001, r=0.455, respectively). p53 expression was not associated with disease-free survival during the 3 years and 9 months long follow-up.
CONCLUSION
A complete disruption of association between p53 and Bak could impair of p53-dependent apoptotic pathway that involves Bak. The relationship of Bak with EPO and EPOR is evidence of their co-expression suggesting competition between EPO mediated cell survival and Bak associated apoptosis in colorectal carcinomas.
Topics: Adenocarcinoma; Cell Nucleus; Colorectal Neoplasms; Cytoplasm; Disease-Free Survival; Erythropoietin; Female; Humans; Immunohistochemistry; Male; Neoplasm Staging; Receptors, Erythropoietin; Tumor Suppressor Protein p53; bcl-2 Homologous Antagonist-Killer Protein
PubMed: 19846965
DOI: No ID Found -
Molecular Cell Sep 2014Apoptotic stimuli activate and oligomerize the proapoptotic proteins Bak and Bax, resulting in mitochondrial outer-membrane permeabilization and subsequent cell death....
Apoptotic stimuli activate and oligomerize the proapoptotic proteins Bak and Bax, resulting in mitochondrial outer-membrane permeabilization and subsequent cell death. This activation can occur when certain BH3-only proteins interact directly with Bak and Bax. Recently published crystal structures reveal that Bax separates into core and latch domains in response to BH3 peptides. The distinguishing characteristics of BH3 peptides capable of directly activating Bax were also elucidated. Here we identify specific BH3 peptides capable of "unlatching" Bak and describe structural insights into Bak activation and oligomerization. Crystal structures and crosslinking experiments demonstrate that Bak undergoes a conformational change similar to that of Bax upon activation. A structure of the Bak core domain dimer provides a high-resolution image of this key intermediate in the pore-forming oligomer. Our results confirm an analogous mechanism for activation and dimerization of Bak and Bax in response to certain BH3 peptides.
Topics: Animals; Crystallography; Cysteine; Humans; Mice; Mitochondria; Models, Molecular; Peptide Fragments; Protein Conformation; Protein Multimerization; Protein Structure, Secondary; Protein Structure, Tertiary; Proto-Oncogene Proteins; bcl-2 Homologous Antagonist-Killer Protein; bcl-2-Associated X Protein
PubMed: 25175025
DOI: 10.1016/j.molcel.2014.07.016 -
Viruses Apr 2023As a zoonotic virus, Japanese Encephalitis virus (JEV) poses a serious threat to human health and the breeding industry. Regarding the mechanism and complications of...
As a zoonotic virus, Japanese Encephalitis virus (JEV) poses a serious threat to human health and the breeding industry. Regarding the mechanism and complications of tissue inflammation caused by JEV, such as encephalitis and orchitis, there is no effective drug treatment currently, and the mechanism of occurrence has not been thoroughly studied. Therefore, it is necessary to study the mechanism of the inflammatory pathway caused by JEV. As one of the key proteins regulating cell death, BCL2 antagonist/killer (BAK) is also a necessary prerequisite for the release of cellular inflammatory factors. We found that after JEV infection, BAK-knockdown cells died less than normal cells, and the transcription levels of inflammatory factors such as , , and and their corresponding regulatory genes were also significantly reduced. By further verifying protein expression on the cell death pathway, it was found that pyroptotic activation and virus titer were also significantly reduced in BAK.KD cells, suggesting that JEV proliferation might be related to BAK-induced cell death. From our data, we could conclude that JEV utilized the BAK-promoted pyroptotic pathway to release more virions after the final Gasdermin D-N (GSDMD-N) protein pore formation for the purpose of JEV proliferation. Therefore, the study of the endogenous cell death activator protein BAK and the final release pathway of JEV, is expected to provide some new theoretical basis for future research on the screening of targeted drugs for the treatment of inflammatory diseases caused by JEV.
Topics: Animals; Humans; Male; Cell Proliferation; Encephalitis Virus, Japanese; Encephalitis Viruses, Japanese; Encephalitis, Japanese; Pyroptosis; Swine; bcl-2 Homologous Antagonist-Killer Protein
PubMed: 37112954
DOI: 10.3390/v15040974 -
BioMed Research International 2014Genetic analyses have provided evidence to suggest that Bax and Bak are the essential genes for apoptosis in mammalians cells. This study aimed to search for biomarkers...
Genetic analyses have provided evidence to suggest that Bax and Bak are the essential genes for apoptosis in mammalians cells. This study aimed to search for biomarkers in breast cancer to be used as prognostic markers for the disease. The Bak and Bax genes expressions were analyzed in 23 breast cancer patients by RT-PCR technique. SSCP technique was used to detect the mobility of the abnormal fragment in Bak exon 4. PCR for Bax promoter was digested with Tau 1 restriction enzyme to identify a single polymorphism G(-248)A. The expression of Bak gene is related to several clinical factors of breast cancer. The analysis of Bax RNA showed 4 isoforms of Bax with different distributions in the normal and tumor tissues. These isoforms were Bax α , d, δ , and ζ . Exon 4 had a normal pattern in all cases of breast cancer. There was a statistically significant difference in the frequency distribution of the G(-248)A genotypes in the breast cancer tissues with grade 3+high, T2 stage, lobular +other, and PR -ve subgroups. In this study, Bak expression seems to lead to development of breast cancer and affects the disease progression. Also, Bax d and Bax δ could be used as risk factor and biomarker for breast cancer with the distribution of G284A.
Topics: Breast Neoplasms; Chi-Square Distribution; Electrophoresis, Agar Gel; Exons; Female; Gene Expression Regulation, Neoplastic; Genetic Predisposition to Disease; Humans; Middle Aged; Mutation; Polymerase Chain Reaction; Polymorphism, Single Nucleotide; Polymorphism, Single-Stranded Conformational; Protein Isoforms; RNA, Messenger; Restriction Mapping; bcl-2 Homologous Antagonist-Killer Protein; bcl-2-Associated X Protein
PubMed: 24672785
DOI: 10.1155/2014/249372 -
ELife Feb 2017During apoptosis, Bak and Bax undergo major conformational change and form symmetric dimers that coalesce to perforate the mitochondrial outer membrane via an unknown...
During apoptosis, Bak and Bax undergo major conformational change and form symmetric dimers that coalesce to perforate the mitochondrial outer membrane via an unknown mechanism. We have employed cysteine labelling and linkage analysis to the full length of Bak in mitochondria. This comprehensive survey showed that in each Bak dimer the N-termini are fully solvent-exposed and mobile, the core is highly structured, and the C-termini are flexible but restrained by their contact with the membrane. Dimer-dimer interactions were more labile than the BH3:groove interaction within dimers, suggesting there is no extensive protein interface between dimers. In addition, linkage in the mobile Bak N-terminus (V61C) specifically quantified association between dimers, allowing mathematical simulations of dimer arrangement. Together, our data show that Bak dimers form disordered clusters to generate lipidic pores. These findings provide a molecular explanation for the observed structural heterogeneity of the apoptotic pore.
Topics: Animals; Apoptosis; Humans; Mice; Mitochondria; Mitochondrial Membranes; Protein Multimerization; bcl-2 Homologous Antagonist-Killer Protein
PubMed: 28182867
DOI: 10.7554/eLife.19944 -
Cell Death & Disease Jul 2019Most antineoplastic chemotherapies eliminate cancer cells through activation of the mitochondria-controlled intrinsic apoptotic pathway. Therein, BAX, BAK, and/or BOK...
Most antineoplastic chemotherapies eliminate cancer cells through activation of the mitochondria-controlled intrinsic apoptotic pathway. Therein, BAX, BAK, and/or BOK function as the essential pore-forming executioners of mitochondrial outer membrane permeabilization (MOMP). The activation threshold of BAX and BAK also correlates inversely with the required strength of an apoptotic stimulus to induce MOMP and thereby effectively determines a cell's readiness to undergo apoptosis. Consequently, the 'gatekeepers' BAX and BAK emerged as therapeutic targets, but functional or genetic loss renders BAX/BAK-targeting strategies prone to fail. Here, we show that the small molecule Raptinal overcomes this limitation by triggering cytochrome c release in a BAX/BAK/BOK-independent manner. Raptinal exerts a dual cytotoxic effect on cancer cells by rapid activation of the intrinsic apoptotic pathway and simultaneous shutdown of mitochondrial function. Together with its efficacy to eliminate cancer cells in vivo, Raptinal could be useful in difficult-to-treat cancer entities harboring defects in the intrinsic apoptosis pathway.
Topics: Apoptosis; Cyclopentanes; Cytochromes c; Fluorenes; HCT116 Cells; Humans; Mitochondria; Mitochondrial Membranes; Neoplasms; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; bcl-2 Homologous Antagonist-Killer Protein; bcl-2-Associated X Protein
PubMed: 31324752
DOI: 10.1038/s41419-019-1790-z