bims-apauto 2022-03-13 papers

bims-apauto

Biomed News

on Apoptosis and autophagy

Issue of 2022–03–13
ten papers selected by
Su Hyun Lee, Seoul National University

How the ends signal the end: Regulation by E3 ubiquitin ligases recognizing protein termini.
Role of autophagy in drug resistance and regulation of osteosarcoma (Review).
Co-targeting of BAX and BCL-XL proteins broadly overcomes resistance to apoptosis in cancer.
BAG family proteins contributes to autophagy-mediated multidrug resistance of tumor.
The V-ATPase complex regulates non-canonical Atg8-family protein lipidation through ATG16L1 recruitment.
Mitochondrial respiration supports autophagy to provide stress resistance during quiescence.
TRAF6 autophagic degradation by avibirnavirus VP3 inhibits antiviral innate immunity via blocking NFKB/NF-κB activation.
KAT7-mediated CANX (calnexin) crotonylation regulates leucine-stimulated MTORC1 activity.
Therapeutic targeting of the USP2-E2F4 axis inhibits autophagic machinery essential for zinc homeostasis in cancer progression.
Incomplete autophagy: Trouble is a friend.

Mol Cell. 2022 Feb 18. pii: S1097-2765(22)00109-5. [Epub ahead of print]

How the ends signal the end: Regulation by E3 ubiquitin ligases recognizing protein termini.

Dawafuti Sherpa, Jakub Chrustowicz, Brenda A Schulman.

  Specificity of eukaryotic protein degradation is determined by E3 ubiquitin ligases and their selective binding to protein motifs, termed "degrons," in substrates for ubiquitin-mediated proteolysis. From the discovery of the first substrate degron and the corresponding E3 to a flurry of recent studies enabled by modern systems and structural methods, it is clear that many regulatory pathways depend on E3s recognizing protein termini. Here, we review the structural basis for recognition of protein termini by E3s and how this recognition underlies biological regulation. Diverse E3s evolved to harness a substrate's N and/or C terminus (and often adjacent residues as well) in a sequence-specific manner. Regulation is achieved through selective activation of E3s and also through generation of degrons at ribosomes or by posttranslational means. Collectively, many E3 interactions with protein N and C termini enable intricate control of protein quality and responses to cellular signals.

Keywords:  C-degron; E3 ligase; GID complex; N-degron; N-end rule; UBR; cullin-RING ligase; protein quality control; ubiquitin

DOI:  https://doi.org/10.1016/j.molcel.2022.02.004
Mol Clin Oncol. 2022 Mar;16(3): 72

Role of autophagy in drug resistance and regulation of osteosarcoma (Review).

Yanchuan Pu, Jin Wang, Shizhong Wang.

  Osteosarcoma (OS) is a malignant tumor originating from primitive mesenchymal tissue that occurs mostly in children and adolescents. It is the most common type of malignant tumor originating from bone. The combination of chemotherapy and surgery is an important treatment strategy for OS; however, multidrug resistance frequently leads to failure of chemotherapy for OS. Autophagy is considered an important mechanism through which bone tumor cells escape apoptosis; inhibition of autophagy may significantly increase the sensitivity of tumor cells to chemotherapeutic drugs. The present review discusses the relationship between chemotherapy resistance and autophagy-related genes, the regulation of autophagy in OS, as well as drugs that inhibit protective autophagy in tumors or cause autophagic death of OS cells or increase their sensitivity to chemotherapy drugs, thereby reducing chemotherapy resistance and increasing efficacy.

Keywords:  autophagy; drug regulation; drug resistance; osteosarcoma

DOI:  https://doi.org/10.3892/mco.2022.2505
Nat Commun. 2022 Mar 07. 13(1): 1199

Co-targeting of BAX and BCL-XL proteins broadly overcomes resistance to apoptosis in cancer.

Andrea Lopez, Denis E Reyna, Nadege Gitego, Felix Kopp, Hua Zhou, Miguel A Miranda-Roman, Lars Ulrik Nordstrøm, Swathi-Rao Narayanagari, Ping Chi, Eduardo Vilar, Aristotelis Tsirigos, Evripidis Gavathiotis.

Deregulation of the BCL-2 family interaction network ensures cancer resistance to apoptosis and is a major challenge to current treatments. Cancer cells commonly evade apoptosis through upregulation of the BCL-2 anti-apoptotic proteins; however, more resistant cancers also downregulate or inactivate pro-apoptotic proteins to suppress apoptosis. Here, we find that apoptosis resistance in a diverse panel of solid and hematological malignancies is mediated by both overexpression of BCL-XL and an unprimed apoptotic state, limiting direct and indirect activation mechanisms of pro-apoptotic BAX. Both survival mechanisms can be overcome by the combination of an orally bioavailable BAX activator, BTSA1.2 with Navitoclax. The combination demonstrates synergistic efficacy in apoptosis-resistant cancer cells, xenografts, and patient-derived tumors while sparing healthy tissues. Additionally, functional assays and genomic markers are identified to predict sensitive tumors to the combination treatment. These findings advance the understanding of apoptosis resistance mechanisms and demonstrate a novel therapeutic strategy for cancer treatment.

DOI: https://doi.org/10.1038/s41467-022-28741-7
Clin Transl Oncol. 2022 Mar 12.

BAG family proteins contributes to autophagy-mediated multidrug resistance of tumor.

Jufang Guo, Xuelian Du, Chaolin Li.

  Multidrug resistance (MDR) is a significant cause of tumor treatment failure. Accumulating evidence suggests that autophagy plays a significant role in the development of MDR. Autophagy is a conserved mechanism that maintains tumor homeostasis by removing damaged mitochondria. However, the specific regulatory mechanism is unclear. Here, we summarize recent studies on the role of autophagy in the development of MDR and the initiation of mitophagy by Bcl-2-associated athanogene (BAG) family proteins. Additionally, this mini-review emphasizes the regulatory role of BAG family proteins, which maintain mitochondrial homeostasis by regulating the PINK1/Parkin pathway. Elucidation of the regulatory mechanisms of mitophagy may foster the development of clinical therapeutic strategies for MDR tumors.

Keywords:  Autophagy; BAG; Mitochondria; Mitophagy; Multidrug resistance

DOI:  https://doi.org/10.1007/s12094-022-02819-6
Autophagy. 2022 Mar 08. 1-2

The V-ATPase complex regulates non-canonical Atg8-family protein lipidation through ATG16L1 recruitment.

Lewis Timimi, Carmen Figueras-Novoa, Elena Marcassa, Oliver Florey, J Kenneth Baillie, Rupert Beale, Rachel Ulferts.

  Conjugation of the Atg8 (autophagy related 8) family of ubiquitin-like proteins to phospholipids of the phagophore is a hallmark of macroautophagy/autophagy. Consequently, Atg8 family members, especially LC3B, are commonly used as a marker of autophagosomes. However, the Atg8 family of proteins are not found solely attached to double-membrane autophagosomes. In non-canonical Atg8-family protein lipidation they become conjugated to single membranes. We have shown that this process is triggered by recruitment of ATG16L1 by the vacuolar-type H+-translocating ATPase (V-ATPase) proton pump, suggesting a role for pH sensing in recruitment of Atg8-family proteins to single membranes.

Keywords:  ATG16L1; Atg4; Atg8; CASM; SopF; V-ATPase; influenza; lipidation; non-canonical autophagy

DOI:  https://doi.org/10.1080/15548627.2022.2029233
Autophagy. 2022 Mar 08. 1-18

Mitochondrial respiration supports autophagy to provide stress resistance during quiescence.

Silvia Magalhaes-Novais, Jan Blecha, Ravindra Naraine, Jana Mikesova, Pavel Abaffy, Alena Pecinova, Mirko Milosevic, Romana Bohuslavova, Jan Prochazka, Shawez Khan, Eliska Novotna, Radek Sindelka, Radek Machan, Mieke Dewerchin, Erik Vlcak, Joanna Kalucka, Sona Stemberkova Hubackova, Ales Benda, Jermaine Goveia, Tomas Mracek, Cyril Barinka, Peter Carmeliet, Jiri Neuzil, Katerina Rohlenova, Jakub Rohlena.

  Mitochondrial oxidative phosphorylation (OXPHOS) generates ATP, but OXPHOS also supports biosynthesis during proliferation. In contrast, the role of OXPHOS during quiescence, beyond ATP production, is not well understood. Using mouse models of inducible OXPHOS deficiency in all cell types or specifically in the vascular endothelium that negligibly relies on OXPHOS-derived ATP, we show that selectively during quiescence OXPHOS provides oxidative stress resistance by supporting macroautophagy/autophagy. Mechanistically, OXPHOS constitutively generates low levels of endogenous ROS that induce autophagy via attenuation of ATG4B activity, which provides protection from ROS insult. Physiologically, the OXPHOS-autophagy system (i) protects healthy tissue from toxicity of ROS-based anticancer therapy, and (ii) provides ROS resistance in the endothelium, ameliorating systemic LPS-induced inflammation as well as inflammatory bowel disease. Hence, cells acquired mitochondria during evolution to profit from oxidative metabolism, but also built in an autophagy-based ROS-induced protective mechanism to guard against oxidative stress associated with OXPHOS function during quiescence.Abbreviations: AMPK: AMP-activated protein kinase; AOX: alternative oxidase; Baf A: bafilomycin A1; CI, respiratory complexes I; DCF-DA: 2',7'-dichlordihydrofluorescein diacetate; DHE: dihydroethidium; DSS: dextran sodium sulfate; ΔΨmi: mitochondrial inner membrane potential; EdU: 5-ethynyl-2'-deoxyuridine; ETC: electron transport chain; FA: formaldehyde; HUVEC; human umbilical cord endothelial cells; IBD: inflammatory bowel disease; LC3B: microtubule associated protein 1 light chain 3 beta; LPS: lipopolysaccharide; MEFs: mouse embryonic fibroblasts; MTORC1: mechanistic target of rapamycin kinase complex 1; mtDNA: mitochondrial DNA; NAC: N-acetyl cysteine; OXPHOS: oxidative phosphorylation; PCs: proliferating cells; PE: phosphatidylethanolamine; PEITC: phenethyl isothiocyanate; QCs: quiescent cells; ROS: reactive oxygen species; PLA2: phospholipase A2, WB: western blot.

Keywords:  ATG4B; biosynthesis; cell death; electron transport chain; endothelial cells; mitochondria; oxidative phosphorylation; oxidative stress; reactive oxygen species

DOI:  https://doi.org/10.1080/15548627.2022.2038898
Autophagy. 2022 Mar 10. 1-18

TRAF6 autophagic degradation by avibirnavirus VP3 inhibits antiviral innate immunity via blocking NFKB/NF-κB activation.

Tingjuan Deng, Boli Hu, Xingbo Wang, Shuxiang Ding, Lulu Lin, Yan Yan, Xiran Peng, Xiaojuan Zheng, Min Liao, Yulan Jin, Weiren Dong, Jinyan Gu, Jiyong Zhou.

  Ubiquitination is an important reversible post-translational modification. Many viruses hijack the host ubiquitin system to enhance self-replication. In the present study, we found that Avibirnavirus VP3 protein was ubiquitinated during infection and supported virus replication by ubiquitination. Mass spectrometry and mutation analysis showed that VP3 was ubiquitinated at residues K73, K135, K158, K193, and K219. Virus rescue showed that ubiquitination at sites K73, K193, and K219 on VP3 could enhance the replication abilities of infectious bursal disease virus (IBDV), and that K135 was essential for virus survival. Binding of the zinc finger domain of TRAF6 (TNF receptor associated factor 6) to VP3 mediated K11- and K33-linked ubiquitination of VP3, which promoted its nuclear accumulation to facilitate virus replication. Additionally, VP3 could inhibit TRAF6-mediated NFKB/NF-κB (nuclear factor kappa B) activation and IFNB/IFN-β (interferon beta) production to evade host innate immunity by inducing TRAF6 autophagic degradation in an SQSTM1/p62 (sequestosome 1)-dependent manner. Our findings demonstrated a macroautophagic/autophagic mechanism by which Avibirnavirus protein VP3 blocked NFKB-mediated IFNB production by targeting TRAF6 during virus infection, and provided a potential drug target for virus infection control.Abbreviations: ATG: autophagy related; BafA1: bafilomycin A1; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; Cas9: CRISPR-associated protein 9; CHX: cycloheximide; Co-IP: co-immunoprecipitation; CRISPR: clustered regularly interspaced short palindromic repeats; GAPDH: glyceraldehyde-3-phosphate dehydrogenase; GST: glutathione S-transferase; IBDV: infectious bursal disease virus; IF: indirect immunofluorescence; IFNB/IFN-β: interferon beta; mAb: monoclonal antibody; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MOI: multiplicity of infection; MS: mass spectrometry; NFKB/NF-κB: nuclear factor kappa B; NBR1: NBR1 autophagy cargo receptor; OPTN: optineurin; pAb: polyclonal antibody; PRRs: pattern recognition receptors; RNF125: ring finger protein 125; RNF135/Riplet: ring finger protein 135; SQSTM1/p62: sequestosome 1; TAX1BP1: tax1 binding protein1; TCID50: 50% tissue culture infective dose; TRAF3: TNF receptor associated factor 3; TRAF6: TNF receptor associated factor 6; TRIM25: tripartite motif containing 25; Ub: ubiquitin; Wort: wortmannin; WT: wild type.

Keywords:  AvibirnavirusVP3 ubiquitination; SQSTM1; TRAF6 degradation; innate antiviral immune; nuclear trafficking; selective autophagy

DOI:  https://doi.org/10.1080/15548627.2022.2047384
Autophagy. 2022 Mar 10. 1-18

KAT7-mediated CANX (calnexin) crotonylation regulates leucine-stimulated MTORC1 activity.

Guokai Yan, Xiuzhi Li, Zilong Zheng, Weihua Gao, Changqing Chen, Xinkai Wang, Zhongyi Cheng, Jie Yu, Geng Zou, Muhammad Zahid Farooq, Xiaoyan Zhu, Weiyun Zhu, Qing Zhong, Xianghua Yan.

  Amino acids play crucial roles in the MTOR (mechanistic target of rapamycin kinase) complex 1 (MTORC1) pathway. However, the underlying mechanisms are not fully understood. Here, we establish a cell-free system to mimic the activation of MTORC1, by which we identify CANX (calnexin) as an essential regulator for leucine-stimulated MTORC1 pathway. CANX translocates to lysosomes after leucine deprivation, and its loss of function renders either the MTORC1 activity or the lysosomal translocation of MTOR insensitive to leucine deprivation. We further find that CANX binds to LAMP2 (lysosomal associated membrane protein 2), and LAMP2 is required for leucine deprivation-induced CANX interaction with the Ragulator to inhibit Ragulator activity toward RRAG GTPases. Moreover, leucine deprivation promotes the lysine (K) 525 crotonylation of CANX, which is another essential condition for the lysosomal translocation of CANX. Finally, we find that KAT7 (lysine acetyltransferase 7) mediates the K525 crotonylation of CANX. Loss of KAT7 renders the MTORC1 insensitivity to leucine deprivation. Our findings provide new insights for the regulatory mechanism of the leucine-stimulated MTORC1 pathway.

Keywords:  CANX; KAT7; LAMP2; MTORC1; leucine; lysine crotonylation; ragulator

DOI:  https://doi.org/10.1080/15548627.2022.2047481
Autophagy. 2022 Mar 06. 1-21

Therapeutic targeting of the USP2-E2F4 axis inhibits autophagic machinery essential for zinc homeostasis in cancer progression.

Wenjing Xiao, Jianqun Wang, Xiaojing Wang, Shuang Cai, Yanhua Guo, Lin Ye, Dan Li, Anpei Hu, Shikai Jin, Boling Yuan, Yi Zhou, Qilan Li, Qiangsong Tong, Liduan Zheng.

  Macroautophagy/autophagy is a conserved cellular process associated with tumorigenesis and aggressiveness, while mechanisms regulating expression of autophagic machinery genes in cancers still remain elusive. Herein, we identified E2F4 (E2F transcription factor 4) as a novel transcriptional activator of cytoprotective autophagy crucial for zinc homeostasis in cancer cells. Gain- and loss-of-function studies showed that E2F4 promoted autophagy in a cell cycle-dependent manner, resulting in facilitated degradation of MT (metallothionein) proteins, elevated distribution of Zn2+ within autophagosomes, decreased labile intracellular zinc ions, and increased growth, invasion, and metastasis of gastric cancer cells. Mechanistically, E2F4 directly regulated the transcription of ATG2A (autophagy related 2A) and ULK2 (unc-51 like autophagy activating kinase 2), leading to autophagic degradation of MT1E, MT1M, and MT1X, while USP2 (ubiquitin specific peptidase 2) stabilized E2F4 protein to induce its transactivation via physical interaction and deubiquitination in cancer cells. Rescue experiments revealed that USP2 harbored oncogenic properties via E2F4-facilitated autophagy and zinc homeostasis. Emetine, a small chemical inhibitor of autophagy, was able to block interaction between UPS2 and E2F4, increase labile intracellular zinc ions, and suppress tumorigenesis and aggressiveness. In clinical gastric cancer specimens, both USP2 and E2F4 were upregulated and associated with poor outcome of patients. These findings indicate that therapeutic targeting of the USP2-E2F4 axis inhibits autophagic machinery essential for zinc homeostasis in cancer progression.Abbreviations: 3-MA: 3-methyladenine; ANOVA: analysis of variance; ATG2A: autophagy related 2A; ATG5: autophagy related 5; ATP: adenosine triphosphate; BECN1: beclin 1; BiFC: bimolecular fluorescence complementation; CCND1: cyclin D1; CDK: cyclin dependent kinase; ChIP: chromatin immunoprecipitation; CHX: cycloheximide; Co-IP: co-immunoprecipitation; DAPI: 4',6-diamidino-2-phenylindole; E2F4: E2F transcription factor 4; eATP: extracellular adenosine triphosphate; EBSS: Earle's balanced salt solution; FP: first progression; FRET: fluorescence resonance energy transfer; FUCCI: fluorescent ubiquitination-based cell cycle indicator; GFP: green fluorescent protein; GST: glutathione S-transferase; HA: hemagglutinin; MAP1LC3B/LC3B: microtubule associated protein 1 light chain 3 beta; MDM2: MDM2 proto-oncogene; MKI67/Ki-67: marker of proliferation Ki-67; MT: metallothionein; MT1E: metallothionein 1E; MT1M: metallothionein 1M; MT1X: metallothionein 1X; MTT: 3-(4,5-dimethyltriazol-2-yl)-2,5-diphenyl tetrazolium bromide; OS: overall survival; PECAM1/CD31: platelet and endothelial cell adhesion molecule 1; PIK3C3: phosphatidylinositol 3-kinase catalytic subunit type 3; qPCR: quantitative PCR; RFP: red fluorescent protein; SQSTM1/p62: sequestosome 1; UBXN1: UBX domain protein 1; Ub: ubiquitin; ULK2: unc-51 like autophagy activating kinase 2; USP14: ubiquitin specific peptidase 14; USP2: ubiquitin specific peptidase 2; USP5: ubiquitin specific peptidase 5; USP7: ubiquitin specific peptidase 7; ZnCl2: zinc chloride.

Keywords:  Autophagy; E2F transcription factor 4; gastric cancer; ubiquitin specific peptidase 2; zinc homeostasis

DOI:  https://doi.org/10.1080/15548627.2022.2044651
Med Res Rev. 2022 Mar 11.

Incomplete autophagy: Trouble is a friend.

Qiang Zhang, Shijie Cao, Feng Qiu, Ning Kang.

  Incomplete autophagy is an impaired self-eating process of intracellular macromolecules and organelles in which accumulated autophagosomes do not fuse with lysosomes for degradation, resulting in the blockage of autophagic flux. In this review, we summarized the literature over the past decade describing incomplete autophagy, and found that different from the double-edged sword effect of general autophagy on promoting cell survival or death, incomplete autophagy plays a crucial role in disrupting cellular homeostasis, and promotes only cell death. What matters is that incomplete autophagy is closely relevant to the pathogenesis and progression of various human diseases, which, meanwhile, intimately linking to the pharmacologic and toxicologic effects of several compounds. Here, we comprehensively reviewed the latest progress of incomplete autophagy on molecular mechanisms and signaling pathways. Moreover, implications of incomplete autophagy for pharmacotherapy are also discussed, which has great relevance for our understanding of the distinctive role of incomplete autophagy in cellular physiology and disease. Consequently, targeting incomplete autophagy may contribute to the development of novel generation therapeutic agents for diverse human diseases.

Keywords:  autophagic flux; incomplete autophagy; molecular mechanisms; pharmacotherapy; toxicology

DOI:  https://doi.org/10.1002/med.21884