scholarly journals Structural Insights into Bak Activation and Oligomerisation

2014 ◽  
Vol 70 (a1) ◽  
pp. C1166-C1166
Author(s):  
Jason Brouwer ◽  
Adeline Robin ◽  
Geoff Thompson ◽  
Ahmad Wardak ◽  
Ruth Kluck ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Cell Death ◽  
Outer Membrane ◽  
Crystal Structures ◽  
Conformational Changes ◽  
Mitochondrial Outer Membrane ◽  
Related Protein ◽  
Mitochondrial Outer Membrane Permeabilisation ◽  
Structural Insights ◽  
Analogous Mechanism

Apoptotic stimuli activate and oligomerise the pro-apoptotic proteins Bak and Bax resulting in mitochondrial outer membrane permeabilisation and subsequent cell death. This activation can occur when certain BH3-only proteins directly interact with Bak and Bax. A recent crystal structure by Czabotar et al. (2013) revealed a novel conformational change for Bax upon activation by BH3-only peptides. Distinguishing characteristics of BH3-only proteins capable of directly activating Bax were also elucidated. Here we describe complementary studies on the related protein Bak. We identify specific BH3-only peptides capable of inducing Bak dimerisation and describe crystal structures that provide key insights into Bak activation and oligomerisation. These structures demonstrate that Bak undergoes similar conformational changes upon activation to those observed with Bax. Altogether our results confirm an analogous mechanism for activation and dimerization of Bak and Bax in response to BH3-only peptides.

scholarly journals Activated BAX/BAK enable mitochondrial inner membrane permeabilisation and mtDNA release during cell death

2018 ◽  
Author(s):  
Joel S Riley ◽  
Giovanni Quarato ◽  
Jonathan Lopez ◽  
Jim O’Prey ◽  
Matthew Pearson ◽  
...  
Keyword(s):  
Cell Death ◽  
Outer Membrane ◽  
Super Resolution ◽  
Mitochondrial Outer Membrane ◽  
Dependent Manner ◽  
Inner Membrane ◽  
Membrane Pores ◽  
Mitochondrial Inner Membrane ◽  
Mitochondrial Outer Membrane Permeabilisation ◽  
Resolution Imaging

AbstractDuring apoptosis, pro-apoptotic BAX and BAK are activated, causing mitochondrial outer membrane permeabilisation (MOMP), caspase activation and cell death. However, even in the absence of caspase activity, cells usually die following MOMP. Such caspase-independent cell death is accompanied by inflammation that requires mitochondrial DNA (mtDNA) activation of cGAS-STING signaling. Because the mitochondrial inner membrane is thought to remain intact during apoptosis, we sought to address how matrix mtDNA could activate the cytosolic cGAS-STING signaling pathway. Strikingly, using super-resolution imaging, we show that mtDNA is efficiently released from mitochondria following MOMP. In a temporal manner, we find that following MOMP, BAX/BAK-mediated mitochondrial outer membrane pores gradually widen over time. This allows extrusion of the mitochondrial inner membrane into the cytosol whereupon it permeablises allowing mtDNA release. Our data demonstrate that mitochondrial inner membrane permeabilisation can occur during cell death in a BAX/BAK-dependent manner. Importantly, by enabling the cytosolic release of mtDNA, inner membrane permeabilisation underpins the immunogenic effects of caspase-independent cell death.

scholarly journals Mutations in Fis1 disrupt orderly disposal of defective mitochondria

2014 ◽  
Vol 25 (1) ◽  
pp. 145-159 ◽  
Author(s):  
Qinfang Shen ◽  
Koji Yamano ◽  
Brian P. Head ◽  
Sumihiro Kawajiri ◽  
Jesmine T. M. Cheung ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Membrane Protein ◽  
Outer Membrane ◽  
Outer Membrane Protein ◽  
Couple Stress ◽  
Mitochondrial Fission ◽  
Mitochondrial Outer Membrane ◽  
Related Protein ◽  
Degradation Processes ◽  
Mitochondrial Toxins

Mitochondrial fission is mediated by the dynamin-related protein Drp1 in metazoans. Drp1 is recruited from the cytosol to mitochondria by the mitochondrial outer membrane protein Mff. A second mitochondrial outer membrane protein, named Fis1, was previously proposed as recruitment factor, but Fis1−/− cells have mild or no mitochondrial fission defects. Here we show that Fis1 is nevertheless part of the mitochondrial fission complex in metazoan cells. During the fission cycle, Drp1 first binds to Mff on the surface of mitochondria, followed by entry into a complex that includes Fis1 and endoplasmic reticulum (ER) proteins at the ER–mitochondrial interface. Mutations in Fis1 do not normally affect fission, but they can disrupt downstream degradation events when specific mitochondrial toxins are used to induce fission. The disruptions caused by mutations in Fis1 lead to an accumulation of large LC3 aggregates. We conclude that Fis1 can act in sequence with Mff at the ER–mitochondrial interface to couple stress-induced mitochondrial fission with downstream degradation processes.

scholarly journals Binding of cardiotonic steroids to Na+,K+-ATPase in the E2P state

2020 ◽  
Vol 118 (1) ◽  
pp. e2020438118
Author(s):  
Ryuta Kanai ◽  
Flemming Cornelius ◽  
Haruo Ogawa ◽  
Kanna Motoyama ◽  
Bente Vilsen ◽  
...  
Keyword(s):  
Crystal Structure ◽  
Crystal Structures ◽  
Conformational Changes ◽  
Ground State ◽  
Sodium Pump ◽  
Structural Features ◽  
Animal Cells ◽  
Inhibitory Potency ◽  
New Members ◽  
Wide Range

The sodium pump (Na+, K+-ATPase, NKA) is vital for animal cells, as it actively maintains Na+ and K+ electrochemical gradients across the cell membrane. It is a target of cardiotonic steroids (CTSs) such as ouabain and digoxin. As CTSs are almost unique strong inhibitors specific to NKA, a wide range of derivatives has been developed for potential therapeutic use. Several crystal structures have been published for NKA-CTS complexes, but they fail to explain the largely different inhibitory properties of the various CTSs. For instance, although CTSs are thought to inhibit ATPase activity by binding to NKA in the E2P state, we do not know if large conformational changes accompany binding, as no crystal structure is available for the E2P state free of CTS. Here, we describe crystal structures of the BeF3− complex of NKA representing the E2P ground state and then eight crystal structures of seven CTSs, including rostafuroxin and istaroxime, two new members under clinical trials, in complex with NKA in the E2P state. The conformations of NKA are virtually identical in all complexes with and without CTSs, showing that CTSs bind to a preformed cavity in NKA. By comparing the inhibitory potency of the CTSs measured under four different conditions, we elucidate how different structural features of the CTSs result in different inhibitory properties. The crystal structures also explain K+-antagonism and suggest a route to isoform specific CTSs.

scholarly journals Poliovirus Induces Bax-Dependent Cell Death Mediated by c-Jun NH2-Terminal Kinase

2007 ◽  
Vol 81 (14) ◽  
pp. 7504-7516 ◽  
Author(s):  
Arnaud Autret ◽  
Sandra Martin-Latil ◽  
Laurence Mousson ◽  
Aurélie Wirotius ◽  
Frédéric Petit ◽  
...  
Keyword(s):  
Cell Death ◽  
Outer Membrane ◽  
Motor Neurons ◽  
Cell Receptor ◽  
Cellular Level ◽  
Membrane Permeabilization ◽  
Paralytic Poliomyelitis ◽  
Receptor Interaction ◽  
Mitochondrial Outer Membrane ◽  
Mitochondrial Outer Membrane Permeabilization

ABSTRACT Poliovirus (PV) is the causal agent of paralytic poliomyelitis, a disease that involves the destruction of motor neurons associated with PV replication. In PV-infected mice, motor neurons die through an apoptotic process. However, mechanisms by which PV induces cell death in neuronal cells remain unclear. Here, we demonstrate that PV infection of neuronal IMR5 cells induces cytochrome c release from mitochondria and loss of mitochondrial transmembrane potential, both of which are evidence of mitochondrial outer membrane permeabilization. PV infection also activates Bax, a proapoptotic member of the Bcl-2 family; this activation involves its conformational change and its redistribution from the cytosol to mitochondria. Neutralization of Bax by vMIA protein expression prevents cytochrome c release, consistent with a contribution of PV-induced Bax activation to mitochondrial outer membrane permeabilization. Interestingly, we also found that c-Jun NH2-terminal kinase (JNK) is activated soon after PV infection and that the PV-cell receptor interaction alone is sufficient to induce JNK activation. Moreover, the pharmacological inhibition of JNK by SP600125 inhibits Bax activation and cytochrome c release. This is, to our knowledge, the first demonstration of JNK-mediated Bax-dependent apoptosis in PV-infected cells. Our findings contribute to our understanding of poliomyelitis pathogenesis at the cellular level.

scholarly journals Gossypol, a BH3 mimetic, induces apoptosis in chronic lymphocytic leukemia cells

Blood ◽  
2008 ◽  
Vol 112 (5) ◽  
pp. 1971-1980 ◽  
Author(s):  
Kumudha Balakrishnan ◽  
William G. Wierda ◽  
Michael J. Keating ◽  
Varsha Gandhi
Keyword(s):  
Cell Death ◽  
Chronic Lymphocytic Leukemia ◽  
Outer Membrane ◽  
Lymphocytic Leukemia ◽  
Mitochondrial Outer Membrane ◽  
Dependent Manner ◽  
Antiapoptotic Proteins ◽  
Downstream Effects ◽  
Mitochondrial Membrane Permeabilization ◽  
Bh3 Mimetic

Abstract Gossypol, a cottonseed extract derivative, acts as a BH3-mimetic, binding to the BH3 pocket of antiapoptotic proteins and displacing pro-death partners to induce apoptosis. However, knowledge on the molecular underpinnings of its downstream effects is limited. Since chronic lymphocytic leukemia (CLL) cells express high levels of antiapoptotic proteins that act as a survival mechanism for these replicationally quiescent lymphocytes, we investigated whether gossypol induces apoptosis in these cells and what mechanism underlies gossypol-mediated cytotoxicity. Gossypol induced cell death in a concentration- and time-dependent manner; 24-hour incubation with 30 μM gossypol resulted in 50% cell death (median; range, 10%-80%; n = 47) that was not abrogated by pan-specific caspase inhibitor. Starting at 4 hours, the mitochondrial outer membrane was significantly permeabilized (median, 77%; range, 54%-93%; n = 15). Mitochondrial outer membrane permeabiliztaion (MOMP) was concurrent with increased production of reactive oxygen species (ROS); however, antioxidants did not abrogate gossypol-induced cell death. Mitochondrial membrane permeabilization was also associated with loss of intracellular adenosine triphosphate (ATP), activation of BAX, and release of cytochrome c and apoptosis-inducing factor (AIF), which was translocated to the nucleus. Blocking AIF translocation resulted in a decreased apoptosis, suggesting that AIF contributes to gossypol-mediated cytotoxicity in CLL lymphocytes.

scholarly journals The mitochondrial outer membrane AAA ATPase AtOM66 affects cell death and pathogen resistance inArabidopsis thaliana

2014 ◽  
Vol 80 (4) ◽  
pp. 709-727 ◽  
Author(s):  
Botao Zhang ◽  
Olivier Van Aken ◽  
Louise Thatcher ◽  
Inge De Clercq ◽  
Owen Duncan ◽  
...  
Keyword(s):  
Cell Death ◽  
Outer Membrane ◽  
Pathogen Resistance ◽  
Mitochondrial Outer Membrane ◽  
Aaa Atpase

scholarly journals Crystal structure of the cyan fluorescent protein Cerulean-S175G

2016 ◽  
Vol 72 (7) ◽  
pp. 516-522 ◽  
Author(s):  
Sang-wook Park ◽  
Sunghyun Kang ◽  
Tae-Sung Yoon
Keyword(s):  
Crystal Structure ◽  
Crystal Structures ◽  
Conformational Changes ◽  
Fluorescent Protein ◽  
Detailed Comparison ◽  
Cyan Fluorescent Protein ◽  
Double Mutation ◽  
Aequorea Victoria ◽  
Green Fluorescent ◽  
Enhanced Cyan Fluorescent Protein

Enhanced cyan fluorescent protein (ECFP) was derived fromAequorea victoriagreen fluorescent protein (avGFP), notably with S65T/Y66W mutations. Its chromophore consists of a tripeptide comprised of Thr65, Trp66 and Gly67 (TWG) residues, while that ofavGFP consists of a Ser65, Tyr66 and Gly67 (SYG) tripeptide. Cerulean and SCFP3A were derived from ECFP-S72A/H148D (a double mutation) with additional Y145A and S175G mutations, respectively, while Cerulean-S175G has both mutations (Y145A and S175G). The crystal structures of these ECFP variants at neutral pH were reported to adopt two distinct major conformations calledECFPandCerulean. In this study, Cerulean-S175G was revealed to adopt only theCeruleanconformation, while Cerulean has been reported to adopt both theECFPand theCeruleanconformations in its crystal structures. Sharing the same S175G mutation with SCFP3A, Cerulean-S175G showed a slightly increased quantum yield, like SCFP3A, but did not adopt theECFPconformation adopted by SCFP3A. Detailed comparison of Cerulean-S175G and other ECFP variants revealed that the notable conformational changes in ECFP variants can be understood mainly in terms of the interaction between the Trp66 residue of the chromophore and residues 145–148 of β-strand 7.

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