bims-proteo Biomed News
on Proteostasis
Issue of 2022‒06‒12
24 papers selected by
Eric Chevet

  1. ACS Chem Biol. 2022 Jun 08.
      The proper trafficking of eukaryotic proteins is essential to cellular function. Genetic, environmental, and other stresses can induce protein mistargeting and, in turn, threaten cellular protein homeostasis. Current methods for measuring protein mistargeting are difficult to translate to living cells, and thus the role of cellular signaling networks in stress-dependent protein mistargeting processes, such as ER pre-emptive quality control (ER pQC), is difficult to parse. Herein, we use genetically encoded peroxidases to characterize protein import into the endoplasmic reticulum (ER). We show that the ERHRP/cytAPEX pair provides good selectivity and sensitivity for both multiplexed protein labeling and for identifying protein mistargeting, using the known ER pQC substrate transthyretin (TTR). Although ERHRP labeling induces formation of detergent-resistant TTR aggregates, this is minimized by using low ERHRP expression, without loss of labeling efficiency. cytAPEX labeling recovers TTR that is mistargeted as a consequence of Sec61 inhibition or ER stress-induced ER pQC. Furthermore, we discover that stress-free activation of the ER stress-associated transcription factor ATF6 recapitulates the TTR import deficiency of ER pQC. Hence, proximity labeling is an effective strategy for characterizing factors that influence ER protein import in living cells.
  2. J Chem Inf Model. 2022 Jun 07.
      Ubiquitination is a type of post-translational modification wherein the small protein ubiquitin (Ub) is covalently bound to a lysine on a target protein. Ubiquitination can signal for several regulatory pathways including protein degradation. Ubiquitination occurs by a series of reactions catalyzed by three types of enzymes: ubiquitin activating enzymes, E1; ubiquitin conjugating enzymes, E2; and ubiquitin ligases, E3. E2 enzymes directly catalyze the transfer of Ub to the target protein─the RING E3 improves the efficiency. Prior to its transfer, Ub is covalently linked to the E2 via a thioester bond and the Ub∼E2 conjugate forms a quaternary complex with the RING E3. It is hypothesized that the RING E3 improves the catalytic efficiency of ubiquitination by placing the E2∼Ub conjugate in a "closed" position, which tensions and weakens the thioester bond. We interrogate this hypothesis by analyzing the strain on the thioester during molecular dynamics simulations of both open and closed E2∼Ub/E3 complexes. Our data indicate that the thioester is strained when the E2∼Ub conjugate is in the closed position. We also show that the amount of strain is consistent with the experimental rate enhancement caused by the RING E3. Finally, our simulations show that the closed configuration increases the populations of key hydrogen bonds in the E2∼Ub active site. This is consistent with another hypothesis stating that the RING E3 enhances reaction rates by preorganizing the substrates.
  3. J Cell Physiol. 2022 Jun 07.
      In insects, the last stage of oogenesis is the process where the chorion layers (eggshell) are synthesized and deposited on the surface of the oocytes by the follicle cells. Protein homeostasis is determined by the fine-tuning of translation and degradation pathways, and the ubiquitin-proteasome system is one of the major degradative routes in eukaryotic cells. The conjugation of ubiquitin to targeted substrates is mediated by the ordered action of E1-activating, E2-conjugating, and E3-ligase enzymes, which covalently link ubiquitin to degradation-targeted proteins delivering them to the proteolytic complex proteasome. Here, we found that the mRNAs encoding polyubiquitin (pUbq), E1, and E2 enzymes are highly expressed in the ovaries of the insect vector of Chagas Disease Rhodnius prolixus. RNAi silencing of pUbq was lethal whereas the silencing of E1 and E2 enzymes resulted in drastic decreases in oviposition and embryo viability. Eggs produced by the E1- and E2-silenced insects presented particular phenotypes of altered chorion ultrastructure observed by high-resolution scanning electron microscopy as well as readings for dityrosine cross-linking and X-ray elemental microanalysis, suggesting a disruption in the secretory routes responsible for the chorion biogenesis. In addition, the ovaries from silenced insects presented altered levels of autophagy-related genes as well as a tendency of upregulation in ER chaperones, indicating a disturbance in the general biosynthetic-secretory pathway. Altogether, we found that E1 and E2 enzymes are essential for chorion biogenesis and that their silencing triggers the modulation of autophagy genes suggesting a coordinated function of both pathways for the progression of choriogenesis.
    Keywords:  E1-activating; E2-conjugating; Rhodnius prolixus; choriogenesis; ubiquitin; vector biology
  4. Essays Biochem. 2022 Jun 09. pii: EBC20210060. [Epub ahead of print]
      Proper regulation of protein homeostasis (proteostasis) is essential for all organisms to survive. A diverse range of post-translational modifications (PTMs) allow precise control of protein abundance, function and cellular localisation. In eukaryotic cells, ubiquitination is a widespread, essential PTM that regulates most, if not all cellular processes. Ubiquitin is added to target proteins via a well-defined enzymatic cascade involving a range of conjugating enzymes and ligases, while its removal is catalysed by a class of enzymes known as deubiquitinases (DUBs). Many human diseases have now been linked to DUB dysfunction, demonstrating the importance of these enzymes in maintaining cellular function. These findings have led to a recent explosion in studying the structure, molecular mechanisms and physiology of DUBs in mammalian systems. Plant DUBs have however remained relatively understudied, with many DUBs identified but their substrates, binding partners and the cellular pathways they regulate only now beginning to emerge. This review focuses on the most recent findings in plant DUB biology, particularly on newly identified DUB substrates and how these offer clues to the wide-ranging roles that DUBs play in the cell. Furthermore, the future outlook on how new technologies in mammalian systems can accelerate the plant DUB field forward is discussed.
    Keywords:  deubiquitinase; plant biology; post translational modification; proteostasis; ubiquitin
  5. EMBO J. 2022 Jun 07. e109777
      Autophagy represents a fundamental mechanism for maintaining cell survival and tissue homeostasis in response to physiological and pathological stress. Autophagy initiation converges on the FIP200-ATG13-ULK1 complex wherein the serine/threonine kinase ULK1 plays a central role. Here, we reveal that the E3 ubiquitin ligase TRIM27 functions as a negative regulatory component of the FIP200-ATG13-ULK1 complex. TRIM27 directly polyubiquitinates ULK1 at K568 and K571 sites with K48-linked ubiquitin chains, with proteasomal turnover maintaining control over basal ULK1 levels. However, during starvation-induced autophagy, TRIM27 catalyzes non-degradative K6- and K11-linked ubiquitination of the serine/threonine kinase 38-like (STK38L) kinase. In turn, STK38L ubiquitination promotes its activation and phosphorylation of ULK1 at Ser495, rendering ULK1 in a permissive state for TRIM27-mediated hyper-ubiquitination of ULK1. This cooperative mechanism serves to restrain the amplitude and duration of autophagy. Further evidence from mouse models shows that basal autophagy levels are increased in Trim27 knockout mice and that Trim27 differentially regulates tumorigenesis and metastasis. Our study identifies a key role of STK38L-TRIM27-ULK1 signaling axis in negatively controlling autophagy with relevance established in human breast cancer.
    Keywords:  STK38L; TRIM27; ULK1; autophagy; tumorigenesis
  6. Cell Stress Chaperones. 2022 Jun 07.
      Heat shock protein 70 (Hsp70) is a molecular chaperone and central regulator of protein homeostasis (proteostasis). Paramount to this role is Hsp70's binding to client proteins and co-chaperones to produce distinct complexes, such that understanding the protein-protein interactions (PPIs) of Hsp70 is foundational to describing its function and dysfunction in disease. Mounting evidence suggests that these PPIs include both "canonical" interactions, which are universally conserved, and "non-canonical" (or "secondary") contacts that seem to have emerged in eukaryotes. These two categories of interactions involve discrete binding surfaces, such that some clients and co-chaperones engage Hsp70 with at least two points of contact. While the contributions of canonical interactions to chaperone function are becoming increasingly clear, it can be challenging to deconvolute the roles of secondary interactions. Here, we review what is known about non-canonical contacts and highlight examples where their contributions have been parsed, giving rise to a model in which Hsp70's secondary contacts are not simply sites of additional avidity but are necessary and sufficient to impart unique functions. From this perspective, we propose that further exploration of non-canonical contacts will generate important insights into the evolution of Hsp70 systems and inspire new approaches for developing small molecules that tune Hsp70-mediated proteostasis.
    Keywords:  Bag domain; Hsp110; J-domain protein; Nucleotide exchange factor; Protein aggregation; Protein folding
  7. Cell Rep. 2022 Jun 07. pii: S2211-1247(22)00713-6. [Epub ahead of print]39(10): 110931
      Coordination of inter-tissue stress signaling is essential for organismal fitness. Neuronal mitochondrial perturbations activate the mitochondrial unfolded-protein response (UPRmt) in the intestine via the mitokine Wnt signaling in Caenorhabditis elegans. Here, we found that the protein disulfide isomerase PDI-6 coordinates inter-tissue UPRmt signaling via regulating the Wnt ligand EGL-20. PDI-6 is expressed in the endoplasmic reticulum (ER) and interacts with EGL-20 through disulfide bonds that are essential for EGL-20 stability and secretion. pdi-6 deficiency results in misfolded EGL-20, which leads to its degradation via ER-associated protein degradation (ERAD) machinery. Expression of PDI-6 declines drastically with aging, and animals with pdi-6 deficiency have decreased lifespan. Overexpression of PDI-6 is sufficient to maintain Wnt/EGL-20 protein levels during aging, activating the UPRmt, and significantly extending lifespan in a Wnt- and UPRmt-dependent manner. Our study reveals that protein disulfide isomerase facilitates Wnt secretion to coordinate the inter-tissue UPRmt signaling and organismal aging.
    Keywords:  CP: Cell biology; PDI; PDI-6; UPR(mt); Wnt signaling; aging; cell-non-autonomous regulation; disulfide bond formation; mitochondrial unfolded-protein response; protein disulfide isomerase
  8. Autophagy. 2022 Jun 05. 1-2
      The protein TRIM5 is under intensive investigation related to its roles in antiviral defense, yet its underlying mechanisms of action remain elusive. In our study, we performed an unbiased identification of TRIM5-interacting partners and found proteins participating in a wide variety of cellular functions. We utilized this proteomics data set to uncover a role for TRIM5 in mitophagy, a mitochondrial quality control system that is impaired in multiple human diseases. Mitochondrial damage triggers the recruitment of TRIM5 to ER-mitochondria contact sites where TRIM5 colocalizes with markers of autophagosome biogenesis. Cells lacking TRIM5 are unable to carry out PRKN-dependent and PRKN-independent mitophagy pathways. TRIM5 knockout cells show reduced mitochondrial function and uncontrolled immune activation in response to mitochondrial damage; phenotypes consistent with a requirement for TRIM5 in mitophagy. Mechanistically, we found that TRIM5 is required for the recruitment of the autophagy initiation machinery to damaged mitochondria, where TRIM5 acts as a scaffold promoting interactions between protein markers of mitochondrial damage and the autophagy initiation machinery.
    Keywords:  APEX2; HIV-1; TRIM5α; autophagy; inflammation; mitochondria; mitophagy; restriction factor; tripartite-motif
  9. Cell Rep. 2022 Jun 07. pii: S2211-1247(22)00695-7. [Epub ahead of print]39(10): 110918
      The proteasome holoenzyme regulates the cellular proteome via degrading most proteins. In its 19-subunit regulatory particle (RP), a heterohexameric ATPase enables protein degradation by injecting protein substrates into the core peptidase. RP assembly utilizes "checkpoints," where multiple dedicated chaperones bind to specific ATPase subunits and control the addition of other subunits. Here, we find that the RP assembly checkpoint relies on two common features of the chaperones. Individual chaperones can distinguish an RP, in which their cognate ATPase persists in the ATP-bound state. Chaperones then together modulate ATPase activity to facilitate RP subunit rearrangements for switching to an active, substrate-processing state in the resulting proteasome holoenzyme. Thus, chaperones may sense ATP binding and hydrolysis as a readout for the quality of the RP complex to generate a functional proteasome holoenzyme. Our findings provide a basis to potentially exploit the assembly checkpoints in situations with known deregulation of proteasomal ATPase chaperones.
    Keywords:  ATPase; CP: Molecular biology; chaperone; not4; proteasome; proteasome storage granule; ubiquitin
  10. J Biol Chem. 2022 Jun 03. pii: S0021-9258(22)00542-7. [Epub ahead of print] 102101
      The heat shock protein 90 (Hsp90) is a molecular chaperone central to client protein folding and maturation in eukaryotic cells. During its chaperone cycle, Hsp90 undergoes ATPase-coupled large-scale conformational changes between open and closed states, where the N-terminal and middle domains of the protein form a compact dimerized conformation. However, the molecular principles of the switching motion between the open and closed states remain poorly understood. Here we show by integrating atomistic and coarse-grained molecular simulations with small-angle X-ray scattering experiments and nuclear magnetic resonance spectroscopy data that Hsp90 exhibits rich conformational dynamics modulated by the charged linker, which connects the N-terminal with the middle domain of the protein. We show that the dissociation of these domains is crucial for the conformational flexibility of the open state, with the separation distance controlled by a beta-sheet motif next to the linker region. Taken together, our results suggest that the conformational ensemble of Hsp90 comprises highly extended states, which could be functionally crucial for client processing.
    Keywords:  MD simulations; chaperone; coarse-grained simulation; heat shock protein (HSP); heat shock protein 90 (Hsp90); molecular dynamics; nuclear magnetic resonance (NMR); small‐angle X‐ray scattering (SAXS)
  11. J Cell Sci. 2022 Jun 01. pii: jcs258997. [Epub ahead of print]135(11):
      The formation of autophagosomes and their fusion with lysosomes are key events that underpin autophagic degradation of cargoes. The core ATG8 system, which consists of the ATG8 family of ubiquitin-like proteins and the machineries that conjugate them onto autophagosomal membranes, are among the most-studied autophagy components. Despite the research focus on the core ATG8 system, there are conflicting reports regarding its essential roles in autophagy. Here, we reconcile prior observations of the core ATG8 system into a unifying model of their function that aims to consider apparently conflicting discoveries. Bypass pathways of autophagy that function independently of the core ATG8 system are also discussed.
    Keywords:  ATG8; Autophagosome; Autophagy; GABARAP; LC3
  12. Elife. 2022 Jun 08. pii: e72780. [Epub ahead of print]11
      Protein methylation occurs predominantly on lysine and arginine residues, but histidine also serves as a methylation substrate. However, a limited number of enzymes responsible for this modification have been reported. Moreover, the biological role of histidine methylation has remained poorly understood to date. Here, we report that human METTL18 is a histidine methyltransferase for the ribosomal protein RPL3 and that the modification specifically slows ribosome traversal on Tyr codons, allowing the proper folding of synthesized proteins. By performing an in vitro methylation assay with a methyl donor analog and quantitative mass spectrometry, we found that His245 of RPL3 is methylated at the τ-N position by METTL18. Structural comparison of the modified and unmodified ribosomes showed stoichiometric modification and suggested a role in translation reactions. Indeed, genome-wide ribosome profiling and an in vitro translation assay revealed that translation elongation at Tyr codons was suppressed by RPL3 methylation. Because the slower elongation provides enough time for nascent protein folding, RPL3 methylation protects cells from the cellular aggregation of Tyr-rich proteins. Our results reveal histidine methylation as an example of a ribosome modification that ensures proteome integrity in cells.
    Keywords:  Methylation; PTM; Ribosome; Ribosome profiling; Translation; biochemistry; chemical biology; chromosomes; gene expression; human; proteostasis
  13. Chem Soc Rev. 2022 Jun 07.
      Proteolysis-targeting chimeras (PROTACs) are heterobifunctional molecules consisting of one ligand that binds to a protein of interest (POI) and another that can recruit an E3 ubiquitin ligase. The chemically-induced proximity between the POI and E3 ligase results in ubiquitination and subsequent degradation of the POI by the ubiquitin-proteasome system (UPS). The event-driven mechanism of action (MOA) of PROTACs offers several advantages compared to traditional occupancy-driven small molecule inhibitors, such as a catalytic nature, reduced dosing and dosing frequency, a more potent and longer-lasting effect, an added layer of selectivity to reduce potential toxicity, efficacy in the face of drug-resistance mechanisms, targeting nonenzymatic functions, and expanded target space. Here, we highlight important milestones and briefly discuss lessons learned about targeted protein degradation (TPD) in recent years and conjecture on the efforts still needed to expand the toolbox for PROTAC discovery to ultimately provide promising therapeutics.
  14. Mol Cell. 2022 Jun 07. pii: S1097-2765(22)00483-X. [Epub ahead of print]
      mRNA function is influenced by modifications that modulate canonical nucleobase behavior. We show that a single modification mediates distinct impacts on mRNA translation in a position-dependent manner. Although cytidine acetylation (ac4C) within protein-coding sequences stimulates translation, ac4C within 5' UTRs impacts protein synthesis at the level of initiation. 5' UTR acetylation promotes initiation at upstream sequences, competitively inhibiting annotated start codons. Acetylation further directly impedes initiation at optimal AUG contexts: ac4C within AUG-flanking Kozak sequences reduced initiation in base-resolved transcriptome-wide HeLa results and in vitro utilizing substrates with site-specific ac4C incorporation. Cryo-EM of mammalian 80S initiation complexes revealed that ac4C in the -1 position adjacent to an AUG start codon disrupts an interaction between C and hypermodified t6A at nucleotide 37 of the initiator tRNA. These findings demonstrate the impact of RNA modifications on nucleobase function at a molecular level and introduce mRNA acetylation as a factor regulating translation in a location-specific manner.
    Keywords:  80S; Kozak; NAT10; ac4C; acetylcytidine; cryo-EM; epitranscriptome; initiation; t6A; translation
  15. Curr Biol. 2022 Jun 06. pii: S0960-9822(22)00658-3. [Epub ahead of print]32(11): R506-R508
      Yuri Shibata and David Komander discuss the composition, regulation and functions of the linear ubiquitin chain assembly complex (LUBAC).
  16. J Am Chem Soc. 2022 Jun 08.
      Sarco/endoplasmic reticulum Ca2+-ATPase (SERCA) is a membrane protein on the endoplasmic reticulum (ER) that transports Ca2+ from the cytosol into the ER. As its function is associated with various biological phenomena, SERCA has been recognized as a promising druggable target. Here, we report the second-strongest SERCA-inhibitory compound known to date, which we isolated from the marine cyanobacterium Leptochromothrix valpauliae and named iezoside (1). The structure of iezoside (1) is fundamentally different from that of any other SERCA inhibitor, and its potency is the strongest among marine natural products (Ki 7.1 nM). In this article, we report our comprehensive analysis of iezoside (1), which covers its isolation, structural characterization supported by density functional theory (DFT) calculations and statistical analysis, total synthesis, and clarification of the mode of action of its potent antiproliferative activity (IC50 6.7 ± 0.4 nM against HeLa cells).
  17. Proc Natl Acad Sci U S A. 2022 Jun 14. 119(24): e2200513119
      Coordinated cell function requires a variety of subcellular organelles to exchange proteins and lipids across physical contacts that are also referred to as membrane contact sites. Such organelle-to-organelle contacts also evoke interest because they can appear in response to metabolic changes, immune activation, and possibly other stimuli. The microscopic size and complex, crowded geometry of these contacts, however, makes them difficult to visualize, manipulate, and understand inside cells. To address this shortcoming, we deposited endoplasmic reticulum (ER)-enriched microsomes purified from rat liver or from cultured cells on a coverslip in the form of a proteinaceous planar membrane. We visualized real-time lipid and protein exchange across contacts that form between this ER-mimicking membrane and lipid droplets (LDs) purified from the liver of rat. The high-throughput imaging possible in this geometry reveals that in vitro LD-ER contacts increase dramatically when the metabolic state is changed by feeding the animal and also when the immune system is activated. Contact formation in both cases requires Rab18 GTPase and phosphatidic acid, thus revealing common molecular targets operative in two very different biological pathways. An optical trap is used to demonstrate physical tethering of individual LDs to the ER-mimicking membrane and to estimate the strength of this tether. These methodologies can potentially be adapted to understand and target abnormal contact formation between different cellular organelles in the context of neurological and metabolic disorders or pathogen infection.
    Keywords:  Rab18; lipid droplets; membrane contact sites; optical trap; supported lipid bilayer
  18. iScience. 2022 Jun 17. 25(6): 104414
      Circulating extracellular vesicles (EVs) contain molecular footprints-lipids, proteins, RNA, and DNA-from their cell of origin. Consequently, EV-associated RNA and proteins have gained widespread interest as liquid-biopsy biomarkers. Yet, an integrative proteo-transcriptomic landscape of EVs and comparison with their cell of origin remains obscure. Here, we report that EVs enrich distinct proteo-transcriptome that does not linearly correlate with their cell of origin. We show that EVs enrich endosomal and extracellular proteins, small RNA (∼13-200 nucleotides) associated with cell differentiation, development, and Wnt signaling. EVs cargo specific RNAs (RNY3, vtRNA, and MIRLET-7) and their complementary proteins (YBX1, IGF2BP2, and SRSF1/2). To ensure an unbiased and independent analyses, we studied 12 cancer cell lines, matching EVs (inhouse and exRNA database), and serum EVs of patients with prostate cancer. Together, we show that EV-RNA-protein complexes may constitute a functional interaction network to protect and regulate molecular access until a function is achieved.
    Keywords:  Cancer systems biology; Microenvironment; Transcriptomics
  19. Cell Death Differ. 2022 Jun 09.
      Activation of oval cells (OCs) has been related to hepatocyte injury during chronic liver diseases including non-alcoholic fatty liver disease (NAFLD). However, OCs plasticity can be affected under pathological environments. We previously found protection against hepatocyte cell death by inhibiting protein tyrosine phosphatase 1B (PTP1B). Herein, we investigated the molecular and cellular processes involved in the lipotoxic susceptibility in OCs expressing or not PTP1B. Palmitic acid (PA) induced apoptotic cell death in wild-type (Ptpn1+/+) OCs in parallel to oxidative stress and impaired autophagy. This lipotoxic effect was attenuated in OCs lacking Ptpn1 that showed upregulated antioxidant defences, increased unfolded protein response (UPR) signaling, higher endoplasmic reticulum (ER) content and elevated stearoyl CoA desaturase (Scd1) expression and activity. These effects in Ptpn1-/- OCs concurred with an active autophagy, higher mitochondrial efficiency and a molecular signature of starvation, favoring lipid droplet (LD) formation and dynamics. Autophagy blockade in Ptpn1-/- OCs reduced Scd1 expression, mitochondrial fitness, LD formation and restored lipoapoptosis, an effect also recapitulated by Scd1 silencing. PTP1B immunostaining was detected in OCs from mouse liver and, importantly, LDs were found in OCs from Ptpn1-/- mice with NAFLD. In conclusion, we demonstrated that Ptpn1 deficiency restrains lipoapoptosis in OCs through a metabolic rewiring towards a "starvation-like" fate, favoring autophagy, mitochondrial fitness and LD formation. Dynamic LD-lysosomal interations likely ensure lipid recycling and, overall, these adaptations protect against lipotoxicity. The identification of LDs in OCs from Ptpn1-/- mice with NAFLD opens therapeutic perspectives to ensure OC viability and plasticity under lipotoxic liver damage.
  20. Cancer Discov. 2022 Jun 08. pii: candisc.1672.2021. [Epub ahead of print]
      Obesity is a global epidemic and a major predisposing factor for cancer. Increasing evidence shows that obesity-associated stress is a key driver of cancer risk and progression. Previous work has identified the phase-separation organelles, stress granules (SGs), as mutant KRAS-dependent mediators of stress adaptation. However, the dependence of tumorigenesis on these organelles is unknown. Here, we establish a causal link between SGs and pancreatic ductal adenocarcinoma (PDAC). Importantly, we uncover that dependence on SGs is drastically heightened in obesity-associated PDAC. Furthermore, we identify a previously unknown regulator and component of SGs, namely the serine/arginine protein kinase 2 (SRPK2), as a specific determinant of SG formation in obesity-associated PDAC. We show that SRPK2-mediated SG formation in obesity-associated PDAC is driven by hyperactivation of the IGF1/PI3K/mTOR/S6K1 pathway, and that S6K1 inhibition selectively attenuates SGs and impairs obesity-associated PDAC development.
  21. Cell Metab. 2022 Jun 01. pii: S1550-4131(22)00187-5. [Epub ahead of print]
      The ovarian-tumor-domain-containing deubiquitinases (OTUDs) block ubiquitin-dependent protein degradation and are involved in diverse signaling pathways. We discovered a rare OTUD3 c.863G>A mutation in a family with an early age of onset of diabetes. This mutation reduces the stability and catalytic activity of OTUD3. We next constructed an experiment with Otud3-/- mice and found that they developed worse obesity, dyslipidemia, and insulin resistance than wild-type mice when challenged with a high-fat diet (HFD). We further found that glucose and fatty acids stimulate CREB-binding-protein-dependent OTUD3 acetylation, promoting its nuclear translocation, where OTUD3 regulates various genes involved in glucose and lipid metabolism and oxidative phosphorylation by stabilizing peroxisome-proliferator-activated receptor delta (PPARδ). Moreover, targeting PPARδ using a specific agonist can partially rescue the phenotype of HFD-fed Otud3-/- mice. We propose that OTUD3 is an important regulator of energy metabolism and that the OTUD3 c.863G>A is associated with obesity and a higher risk of diabetes.
    Keywords:  OTUD3; PPARδ; acetylation; metabolic homeostasis; nuclear translocation
  22. EMBO Mol Med. 2022 Jun 07. e15851
      Aberrant localization of proteins to mitochondria disturbs mitochondrial function and contributes to the pathogenesis of Huntington's disease (HD). However, the crucial factors and the molecular mechanisms remain elusive. Here, we found that heat shock transcription factor 1 (HSF1) accumulates in the mitochondria of HD cell models, a YAC128 mouse model, and human striatal organoids derived from HD induced pluripotent stem cells (iPSCs). Overexpression of mitochondria-targeting HSF1 (mtHSF1) in the striatum causes neurodegeneration and HD-like behavior in mice. Mechanistically, mtHSF1 facilitates mitochondrial fission by activating dynamin-related protein 1 (Drp1) phosphorylation at S616. Moreover, mtHSF1 suppresses single-stranded DNA-binding protein 1 (SSBP1) oligomer formation, which results in mitochondrial DNA (mtDNA) deletion. The suppression of HSF1 mitochondrial localization by DH1, a unique peptide inhibitor, abolishes HSF1-induced mitochondrial abnormalities and ameliorates deficits in an HD animal model and human striatal organoids. Altogether, our findings describe an unsuspected role of HSF1 in contributing to mitochondrial dysfunction, which may provide a promising therapeutic target for HD.
    Keywords:  Huntington's disease; heat shock transcription factor 1; human striatal organoids; mitochondrial DNA; single-stranded DNA-binding protein 1
  23. Annu Rev Genet. 2022 Jun 09.
      Autophagy, a lysosome-mediated degradation process evolutionarily conserved from yeast to mammals, is essential for maintaining cellular homeostasis and combating diverse cellular stresses. Autophagy involves de novo synthesis of a double-membrane autophagosome, sequestration of selected cellular contents, and subsequent delivery of sequestrated contents to the vacuole (in yeasts and plants) or to lysosomes (in animal cells) for degradation and recycling. Genetic studies in unicellular and multicellular model organisms have systematically revealed the molecular machinery, regulation, and function of autophagy in physiological settings. I review genetic studies in model organisms-from yeast to worm to fly-that enable us to not only identify autophagy genes, including ATG genes and the metazoan-specific EPG genes, but also uncover variants of autophagy in developmental contexts, novel regulatory mechanisms, and signaling events involved in mediating systemic autophagy response. Genetic analysis also helps us understand the liquid-liquid phase separation and transition that control autophagic degradation of protein aggregates. The emerging role of autophagy in zebrafish tissue regeneration is also discussed. Expected final online publication date for the Annual Review of Genetics, Volume 56 is November 2022. Please see for revised estimates.
  24. Cell. 2022 Jun 01. pii: S0092-8674(22)00593-1. [Epub ahead of print]
      Hijacking the cellular protein degradation system offers unique opportunities for drug discovery, as exemplified by proteolysis-targeting chimeras. Despite their great promise for medical chemistry, so far, it has not been possible to reprogram the bacterial degradation machinery to interfere with microbial infections. Here, we develop small-molecule degraders, so-called BacPROTACs, that bind to the substrate receptor of the ClpC:ClpP protease, priming neo-substrates for degradation. In addition to their targeting function, BacPROTACs activate ClpC, transforming the resting unfoldase into its functional state. The induced higher-order oligomer was visualized by cryo-EM analysis, providing a structural snapshot of activated ClpC unfolding a protein substrate. Finally, drug susceptibility and degradation assays performed in mycobacteria demonstrate in vivo activity of BacPROTACs, allowing selective targeting of endogenous proteins via fusion to an established degron. In addition to guiding antibiotic discovery, the BacPROTAC technology presents a versatile research tool enabling the inducible degradation of bacterial proteins.
    Keywords:  AAA proteases; antimicrobials; bacterial PROTACs; targeted protein degradation