bims-apauto 2021-08-15 papers

Issue of 2021–08–15
six papers selected by
Su Hyun Lee, Seoul National University

Proc Natl Acad Sci U S A. 2021 Aug 17. pii: e2107321118. [Epub ahead of print]118(33):

p62-containing, proteolytically active nuclear condensates, increase the efficiency of the ubiquitin-proteasome system.

Afu Fu, Victoria Cohen-Kaplan, Noa Avni, Ido Livneh, Aaron Ciechanover.

Degradation of a protein by the ubiquitin-proteasome system (UPS) is a multistep process catalyzed by sequential reactions. Initially, ubiquitin is conjugated to the substrate in a process mediated by concerted activity of three enzymes; the last of them-a ubiquitin ligase (E3)-belongs to a family of several hundred members, each recognizing a few specific substrates. This is followed by repeated addition of ubiquitin moieties to the previously conjugated one to generate a ubiquitin chain that serves as a recognition element for the proteasome, which then degrades the substrate. Ubiquitin is recycled via the activity of deubiquitinating enzymes (DUBs). It stands to reason that efficiency of such a complex process would depend on colocalization of the different components in an assembly that allows the reactions to be carried out sequentially and processively. Here we describe nuclear condensates that are dynamic in their composition. They contain p62 as an essential component. These assemblies are generated by liquid-liquid phase separation (LLPS) and also contain ubiquitinated targets, 26S proteasome, the three conjugating enzymes, and DUBs. Under basal conditions, they serve as efficient centers for proteolysis of nuclear proteins (e.g., c-Myc) and unassembled subunits of the proteasome, suggesting they are involved in cellular protein quality control. Supporting this notion is the finding that such foci are also involved in degradation of misfolded proteins induced by heat and oxidative stresses, following recruitment of heat shock proteins and their associated ubiquitin ligase CHIP.

Keywords: LLPS condensates; p62; proteasome; protein degradation; ubiquitin

DOI: https://doi.org/10.1073/pnas.2107321118

Autophagy. 2021 Aug 12. 1-16

TRIM44 links the UPS to SQSTM1/p62-dependent aggrephagy and removing misfolded proteins.

Lin Lyu, Zheng Chen, Nami McCarty.

Until recently, the ubiquitin-proteasome system (UPS) and macroautophagy/autophagy were considered to be two independent systems that target proteins for degradation by proteasomes or via lysosomes, respectively. Here, we report that TRIM44 (tripartite motif containing 44) is a novel link that connects the UPS system with the autophagy degradation pathway. Suppressing the UPS degradation pathway leads to TRIM44 upregulation, which further promotes aggregated protein clearance through the binding of K48 ubiquitin chains on proteins. TRIM44 expression activates autophagy via promoting SQSTM1/p62 oligomerization, which rapidly increases the rate of aggregate protein removal. Overall, our data reveal that TRIM44 is a newly identified link between the UPS system and the autophagy pathway. Delineating the cross-talk between these two degradation pathways may reveal new mechanisms of targeting aggregate-prone diseases, such as cancer and neurodegenerative disease.Abbreviations: 3-MA: 3-methyladenine; ACTB: actin beta; ATG5: autophagy related 5; BB: B-box domain; BECN1: beclin1; BM: bone marrow; CC: coiled-coil domain; CFTR: cystic fibrosis transmembrane conductance regulator; CON: control; CQ: chloroquine; DOX: doxycycline; DSP: dithiobis(succinimidly propionate); ER: endoplasmic reticulum; FI: fluorescence intensity; FL: full length; HIF1A/HIF-1#x3B1;: hypoxia inducible factor 1 subunit alpha; HSC: hematopoietic stem cells; HTT: huntingtin; KD: knockdown; KD-CON: knockdown construct control; MM: multiple myeloma; MTOR: mechanistic target of rapamycin kinase; NP-40: nonidet P-40; NFE2L2/NRF2: nuclear factor, erythroid 2 like 2; OE: overexpression; OE-CON: overexpression construct control; PARP: poly (ADP-ribose) polymerase; SDS: sodium dodecyl sulfate; SQSTM1/p62: sequestosome 1; Tet-on: tetracycline; TRIM44: tripartite motif containing 44; UPS: ubiquitin-proteasome system; ZF: zinc-finger.

Keywords: Aggregates; TRIM44; autophagy; deubiquitinase; misfolded proteins; protein homeostasis; ubiquitin-proteasome system

DOI: https://doi.org/10.1080/15548627.2021.1956105

Autophagy. 2021 Aug 12. 1-2

AMPK-activated ULK1 phosphorylates PIKFYVE to drive formation of PtdIns5P-containing autophagosomes during glucose starvation.

Cansu Karabiyik, David C Rubinsztein.

The induction of macroautophagy/autophagy upon glucose deprivation can occur independently of the PIK3C3/VPS34 complex. Recently, we described a non-canonical signaling pathway involving the kinases AMPK, ULK1 and PIKFYVE that are induced during glucose starvation, leading to the formation of PtdIns5P-containing autophagosomes, resulting in increased autophagy flux and clearance of autophagy substrates. In this cascade, the activation of AMPK leads to ULK1 phosphorylation. ULK1 then phosphorylates PIKFYVE at S1548, leading to its activation and increased PtdIns5P formation, which enables the recruitment of machinery required for autophagosome biogenesis.

Keywords: AMPK; Autophagy; PIKFYVE; Ptdins5P; ULK1; glucose starvation

DOI: https://doi.org/10.1080/15548627.2021.1961409

Autophagy. 2021 Aug 12. 1-2

MTORC1-dependent crinophagy regulates glucagon content in pancreatic α-cells.

Sangam Rajak, Paul M Yen, Rohit A Sinha.

Hormone synthesis and secretion is a highly regulated process governed by metabolic cues. Although peptide hormone action is largely governed by the rate of its synthesis and secretion by endocrine cells, and the levels of its receptors on the target cells, intracellular degradation of the hormone-containing secretory vesicles by lysosomes (crinophagy) adds an additional layer of regulation. In our recent study, we uncovered the regulatory mechanism governing the crinophagic turnover of GCG (glucagon), a glycoprotein hormone secreted by pancreatic α-cells. Our results showed that inhibition of MTORC1 induces crinophagy-mediated degradation of glucagon and decreases its secretion in response to hypoglycemia. Furthermore, we demonstrated that crinophagy-regulated glucagon turnover does not involve macroautophagy. These results suggest that modulation of crinophagy may serve as a novel therapeutic strategy to regulate hormone secretion in endocrine and metabolic pathologies.

Keywords: Autophagy; MTORC1; crinophagy; diabetes; glucagon; lysosomes; rapamycin

DOI: https://doi.org/10.1080/15548627.2021.1961074

Nat Commun. 2021 08 10. 12(1): 4814

Two parallel pathways connect glutamine metabolism and mTORC1 activity to regulate glutamoptosis.

Clément Bodineau, Mercedes Tomé, Sarah Courtois, Ana S H Costa, Marco Sciacovelli, Benoit Rousseau, Elodie Richard, Pierre Vacher, Carlos Parejo-Pérez, Emilie Bessede, Christine Varon, Pierre Soubeyran, Christian Frezza, Piedad Del Socorro Murdoch, Victor H Villar, Raúl V Durán.

Glutamoptosis is the induction of apoptotic cell death as a consequence of the aberrant activation of glutaminolysis and mTORC1 signaling during nutritional imbalance in proliferating cells. The role of the bioenergetic sensor AMPK during glutamoptosis is not defined yet. Here, we show that AMPK reactivation blocks both the glutamine-dependent activation of mTORC1 and glutamoptosis in vitro and in vivo. We also show that glutamine is used for asparagine synthesis and the GABA shunt to produce ATP and to inhibit AMPK, independently of glutaminolysis. Overall, our results indicate that glutamine metabolism is connected with mTORC1 activation through two parallel pathways: an acute alpha-ketoglutarate-dependent pathway; and a secondary ATP/AMPK-dependent pathway. This dual metabolic connection between glutamine and mTORC1 must be considered for the future design of therapeutic strategies to prevent cell growth in diseases such as cancer.

DOI: https://doi.org/10.1038/s41467-021-25079-4

Autophagy. 2021 Aug 12. 1-17

Lipotoxicity-induced STING1 activation stimulates MTORC1 and restricts hepatic lipophagy.

Kunpeng Liu, Dongbo Qiu, Xue Liang, Yingqi Huang, Yao Wang, Xin Jia, Kun Li, Jingyuan Zhao, Cong Du, Xiusheng Qiu, Jun Cui, Zhendong Xiao, Yunfei Qin, Qi Zhang.

Lipid accumulation often leads to lipotoxic injuries to hepatocytes, which can cause nonalcoholic steatohepatitis. The association of inflammation with lipid accumulation in liver tissue has been studied for decades; however, key mechanisms have been identified only recently. In particular, it is still unknown how hepatic inflammation regulates lipid metabolism in hepatocytes. Herein, we found that PA treatment or direct stimulation of STING1 promoted, whereas STING1 deficiency impaired, MTORC1 activation, suggesting that STING1 is involved in PA-induced MTORC1 activation. Mechanistic studies revealed that STING1 interacted with several components of the MTORC1 complex and played an important role in the complex formation of MTORC1 under PA treatment. The involvement of STING1 in MTORC1 activation was dependent on SQSTM1, a key regulator of the MTORC1 pathway. In SQSTM1-deficient cells, the interaction of STING1 with the components of MTORC1 was weak. Furthermore, the impaired activity of MTORC1 via rapamycin treatment or STING1 deficiency decreased the numbers of LDs in cells. PA treatment inhibited lipophagy, which was not observed in STING1-deficient cells or rapamycin-treated cells. Restoration of MTORC1 activity via treatment with amino acids blocked lipophagy and LDs degradation. Finally, increased MTORC1 activation concomitant with STING1 activation was observed in liver tissues of nonalcoholic fatty liver disease patients, which provided clinical evidence for the involvement of STING1 in MTORC1 activation. In summary, we identified a novel regulatory loop of STING1-MTORC1 and explain how hepatic inflammation regulates lipid accumulation. Our findings may facilitate the development of new strategies for clinical treatment of hepatic steatosis.Abbreviations: AA: amino acid; ACTB: actin beta; cGAMP: cyclic GMP-AMP; CGAS: cyclic GMP-AMP synthase; DEPTOR: DEP domain containing MTOR interacting protein; EIF4EBP1: eukaryotic translation initiation factor 4E binding protein 1; FFAs: free fatty acids; GFP: green fluorescent protein; HFD: high-fat diet; HT-DNA: herring testis DNA; IL1B: interleukin 1 beta; LAMP1: lysosomal associated membrane protein 1; LDs: lipid droplets; MAP1LC3: microtubule associated protein 1 light chain 3; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; MEFs: mouse embryonic fibroblasts; MLST8: MTOR associated protein, LST8 homolog; MT-ND1: mitochondrially encoded NADH: ubiquinone oxidoreductase core subunit 1; mtDNA: mitochondrial DNA; MTOR: mechanistic target of rapamycin kinase; MTORC1: MTOR complex 1; NAFL: nonalcoholic fatty liver; NAFLD: nonalcoholic fatty liver disease; NASH: nonalcoholic steatohepatitis; NPCs: non-parenchymal cells; PA: palmitic acid; PLIN2: perilipin 2; RD: regular diet; RELA: RELA proto-oncogene, NF-kB subunit; RPS6: ribosomal protein S6; RPS6KB1: ribosomal protein S6 kinase B1; RPTOR: regulatory associated protein of MTOR complex 1; RRAGA: Ras related GTP binding A; RRAGC: Ras related GTP binding C; SQSTM1: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TBK1: TANK binding kinase 1; TGs: triglycerides; TREX1: three prime repair exonuclease 1.

Keywords: Lipophagy; MTORC1; NAFLD; STING1; TBK1

DOI: https://doi.org/10.1080/15548627.2021.1961072