bims-proteo Biomed News
on Proteostasis
Issue of 2024–03–10
25 papers selected by
Eric Chevet, INSERM
  1. The derlin Dfm1 couples retrotranslocation of a folded protein domain to its proteasomal degradation.
  2. Integrated stress response signaling acts as a metabolic sensor in fat tissues to regulate oocyte maturation and ovulation.
  3. Loss of Grp170 results in catastrophic disruption of endoplasmic reticulum function.
  4. Identification of molecular signatures defines the differential proteostasis response in induced spinal and cranial motor neurons.
  5. Pink1 balances reticulophagy and mitophagy by regulating distinct E3 ubiquitin ligases.
  6. AGO1 controls protein folding in mouse embryonic stem cell fate decisions.
  7. Mechanistic insights into the interactions of TAX1BP1 with RB1CC1 and mammalian ATG8 family proteins.
  8. Proteostatic tuning underpins the evolution of novel multicellular traits.
  9. Tight regulation of a nuclear HAPSTR1-HUWE1 pathway essential for mammalian life.
  10. International consensus guidelines for the definition, detection, and interpretation of autophagy-dependent ferroptosis.
  11. Mapping the substrate landscape of protein phosphatase 2A catalytic subunit PPP2CA.
  12. Oncogenic mutations of KRAS modulate its turnover by the CUL3/LZTR1 E3 ligase complex.
  13. Evidence that Xrn1 is in complex with Gcn1, and is required for full levels of eIF2α phosphorylation.
  14. Loss of WIPI4 in neurodegeneration causes autophagy-independent ferroptosis.
  15. RHBDL4-triggered downregulation of COPII adaptor protein TMED7 suppresses TLR4-mediated inflammatory signaling.
  16. Beyond microtubules: The cellular environment at the endoplasmic reticulum attracts proteins to the nucleus, enabling nuclear transport.
  17. Identification of the YIPF3-YIPF4 heterodimer as a novel Golgiphagy receptor.
  18. The ER tether VAPA is required for proper cell motility and anchors ER-PM contact sites to focal adhesions.
  19. Deficiency of ASGR1 promotes liver injury by increasing GP73-mediated hepatic endoplasmic reticulum stress.
  20. Riboformer: a deep learning framework for predicting context-dependent translation dynamics.
  21. Structural dynamics of RAF1-HSP90-CDC37 and HSP90 complexes reveal asymmetric client interactions and key structural elements.
  22. Basement membrane diversification relies on two competitive secretory routes defined by Rab10 and Rab8 and modulated by dystrophin and the exocyst complex.
  23. Generalized biomolecular modeling and design with RoseTTAFold All-Atom.
  24. Pharmacological induction of autophagy reduces inflammation in macrophages by degrading immunoproteasome subunits.
  25. Molecular mechanism of ATF6 in unfolded protein response and its role in disease.
  1. J Cell Biol. 2024 May 06. pii: e202308074. [Epub ahead of print]223(5):
    The derlin Dfm1 couples retrotranslocation of a folded protein domain to its proteasomal degradation.
    Daniela G Vitali, Daniel Fonseca, Pedro Carvalho.
      Endoplasmic reticulum (ER) proteins are degraded by proteasomes in the cytosol through ER-associated degradation (ERAD). This process involves the retrotranslocation of substrates across the ER membrane, their ubiquitination, and membrane extraction by the Cdc48/Npl4/Ufd1 ATPase complex prior to delivery to proteasomes for degradation. How the presence of a folded luminal domain affects substrate retrotranslocation and this event is coordinated with subsequent ERAD steps remains unknown. Here, using a model substrate with a folded luminal domain, we showed that Cdc48 ATPase activity is sufficient to drive substrate retrotranslocation independently of ERAD membrane components. However, the complete degradation of the folded luminal domain required substrate-tight coupling of retrotranslocation and proteasomal degradation, which was ensured by the derlin Dfm1. Mutations in Dfm1 intramembrane rhomboid-like or cytosolic Cdc48-binding regions resulted in partial degradation of the substrate with accumulation of its folded domain. Our study revealed Dfm1 as a critical regulator of Cdc48-driven retrotranslocation and highlights the importance of coordinating substrate retrotranslocation and degradation during ERAD.
    DOI:  https://doi.org/10.1083/jcb.202308074
  2. Cell Rep. 2024 Mar 06. pii: S2211-1247(24)00191-8. [Epub ahead of print]43(3): 113863
    Integrated stress response signaling acts as a metabolic sensor in fat tissues to regulate oocyte maturation and ovulation.
    Lydia Grmai, Manuel Michaca, Emily Lackner, Narayanan Nampoothiri V P, Deepika Vasudevan.
      Reproduction is an energy-intensive process requiring systemic coordination. However, the inter-organ signaling mechanisms that relay nutrient status to modulate reproductive output are poorly understood. Here, we use Drosophila melanogaster as a model to establish the integrated stress response (ISR) transcription factor, Atf4, as a fat tissue metabolic sensor that instructs oogenesis. We demonstrate that Atf4 regulates lipase activity to mediate yolk lipoprotein synthesis in the fat body. Depletion of Atf4 in the fat body also blunts oogenesis recovery after amino acid deprivation and re-feeding, suggestive of a nutrient-sensing role for Atf4. We also discovered that Atf4 promotes secretion of a fat-body-derived neuropeptide, CNMamide, which modulates neural circuits that promote egg-laying behavior (ovulation). Thus, we posit that ISR signaling in fat tissue acts as a "metabolic sensor" that instructs female reproduction-directly by impacting yolk lipoprotein production and follicle maturation and systemically by regulating ovulation.
    Keywords:  Atf4; CNMa; CP: Cell biology; CP: Developmental biology; adipocyte; bmm; egg retention; integrated stress response; oogenesis; ovulation; yolk protein
    DOI:  https://doi.org/10.1016/j.celrep.2024.113863
  3. Mol Biol Cell. 2024 Mar 06. mbcE24010012
    Loss of Grp170 results in catastrophic disruption of endoplasmic reticulum function.
    Melissa J Mann, Chris Melendez-Suchi, Hannah E Vorndran, Maria Sukhoplyasova, Ashley R Flory, Mary Carson Irvine, Anuradha R Iyer, Christopher J Guerriero, Jeffrey L Brodsky, Linda M Hendershot, Teresa M Buck.
      GRP170 (Hyou1) is required for mouse embryonic development, and its ablation in kidney nephrons leads to renal failure. Unlike most chaperones, GRP170 is the lone member of its chaperone family in the ER lumen. However, the cellular requirement for GRP170, which both binds non-native proteins and acts as nucleotide exchange factor for BiP, is poorly understood. Here, we report on the isolation of mouse embryonic fibroblasts obtained from mice in which LoxP sites were engineered in the Hyou1 loci (Hyou1LoxP/LoxP). A doxycycline-regulated Cre recombinase was stably introduced into these cells. Induction of Cre resulted in depletion of Grp170 protein which culminated in cell death. As Grp170 levels fell we observed a portion of BiP fractionating with insoluble material, increased binding of BiP to a client with a concomitant reduction in its turnover, and reduced solubility of an aggregation-prone BiP substrate. Consistent with disrupted BiP functions, we observed reactivation of BiP and induction of the unfolded protein response (UPR) in futile attempts to provide compensatory increases in ER chaperones and folding enzymes. Together, these results provide insights into the cellular consequences of controlled Grp170 loss and provide hypotheses as to why mutations in the Hyou1 locus are linked to human disease.
    DOI:  https://doi.org/10.1091/mbc.E24-01-0012
  4. Cell Rep. 2024 Mar 07. pii: S2211-1247(24)00213-4. [Epub ahead of print]43(3): 113885
    Identification of molecular signatures defines the differential proteostasis response in induced spinal and cranial motor neurons.
    Ana Paula Zen Petisco Fiore, Shuvadeep Maity, Lauren Jeffery, Disi An, Justin Rendleman, Dylan Iannitelli, Hyungwon Choi, Esteban Mazzoni, Christine Vogel.
      Amyotrophic lateral sclerosis damages proteostasis, affecting spinal and upper motor neurons earlier than a subset of cranial motor neurons. To aid disease understanding, we exposed induced cranial and spinal motor neurons (iCrMNs and iSpMNs) to proteotoxic stress, under which iCrMNs showed superior survival, quantifying the transcriptome and proteome for >8,200 genes at 0, 12, and 36 h. Two-thirds of the proteome showed cell-type differences. iSpMN-enriched proteins related to DNA/RNA metabolism, and iCrMN-enriched proteins acted in the endoplasmic reticulum (ER)/ER chaperone complex, tRNA aminoacylation, mitochondria, and the plasma/synaptic membrane, suggesting that iCrMNs expressed higher levels of proteins supporting proteostasis and neuronal function. When investigating the increased proteasome levels in iCrMNs, we showed that the activity of the 26S proteasome, but not of the 20S proteasome, was higher in iCrMNs than in iSpMNs, even after a stress-induced decrease. We identified Ublcp1 as an iCrMN-specific regulator of the nuclear 26S activity.
    Keywords:  CP: Cell biology; CP: Neuroscience; Ublcp1; amyotrophic lateral sclerosis; motor neurons; proteasome; unfolded protein response
    DOI:  https://doi.org/10.1016/j.celrep.2024.113885
  5. Autophagy. 2024 Mar 08. 1-2
    Pink1 balances reticulophagy and mitophagy by regulating distinct E3 ubiquitin ligases.
    Zhangyuan Yin, Daniel J Klionsky.
      The selective clearance of unwanted, damaged or dangerous components by macroautophagy/autophagy is critical for maintaining cellular homeostasis in organisms from yeast to humans. In recent years, significant progress has been made in understanding how phagophores selectively sequester specific cargo. Nevertheless, a fundamental question remains: Can distinct selective autophagy programs simultaneously operate within the same cell? A recent study from the Baehrecke lab has unveiled a developmentally programmed Pink1-dependent reticulophagy process in the Drosophila intestine. Furthermore, this study demonstrated that autophagy differentially clears mitochondria and ER in the same cell under the regulation of Pink1 through different E3 ubiquitin ligases, highlighting the need for further exploration in understanding the complexity of autophagic substrate selection and crosstalk between diverse autophagy programs.
    Keywords:  Autophagy receptor; Keap1; Pink1; mitophagy; selective autophagy
    DOI:  https://doi.org/10.1080/15548627.2024.2323294
  6. Dev Cell. 2024 Mar 01. pii: S1534-5807(24)00104-7. [Epub ahead of print]
    AGO1 controls protein folding in mouse embryonic stem cell fate decisions.
    Qiuying Liu, Rachel M Pepin, Mariah K Novak, Katharine R Maschhoff, Kailey Worner, Wenqian Hu.
      Argonaute (AGO) proteins are evolutionarily conserved RNA-binding proteins that control gene expression through the small RNAs they interact with. Whether AGOs have regulatory roles independent of RNAs, however, is unknown. Here, we show that AGO1 controls cell fate decisions through facilitating protein folding. We found that in mouse embryonic stem cells (mESCs), while AGO2 facilitates differentiation via the microRNA (miRNA) pathway, AGO1 controls stemness independently of its binding to small RNAs. We determined that AGO1 specifically interacts with HOP, a co-chaperone for the HSP70 and HSP90 chaperones, and enhances the folding of a set of HOP client proteins with intrinsically disordered regions. This AGO1-mediated facilitation of protein folding is important for maintaining stemness in mESCs. Our results demonstrate divergent functions between AGO1 and AGO2 in controlling cellular states and identify an RNA-independent function of AGO1 in controlling gene expression and cell fate decisions.
    Keywords:  AGO1; AGO2; HOP; cell fate decisions; protein folding
    DOI:  https://doi.org/10.1016/j.devcel.2024.02.006
  7. Proc Natl Acad Sci U S A. 2024 Mar 12. 121(11): e2315550121
    Mechanistic insights into the interactions of TAX1BP1 with RB1CC1 and mammalian ATG8 family proteins.
    Mingfang Zhang, Yingli Wang, Xinyu Gong, Yaru Wang, Yuchao Zhang, Yubin Tang, Xindi Zhou, Haobo Liu, Yichao Huang, Jing Zhang, Lifeng Pan.
      TAX1BP1, a multifunctional autophagy adaptor, plays critical roles in different autophagy processes. As an autophagy receptor, TAX1BP1 can interact with RB1CC1, NAP1, and mammalian ATG8 family proteins to drive selective autophagy for relevant substrates. However, the mechanistic bases underpinning the specific interactions of TAX1BP1 with RB1CC1 and mammalian ATG8 family proteins remain elusive. Here, we find that there are two distinct binding sites between TAX1BP1 and RB1CC1. In addition to the previously reported TAX1BP1 SKICH (skeletal muscle and kidney enriched inositol phosphatase (SKIP) carboxyl homology)/RB1CC1 coiled-coil interaction, the first coiled-coil domain of TAX1BP1 can directly bind to the extreme C-terminal coiled-coil and Claw region of RB1CC1. We determine the crystal structure of the TAX1BP1 SKICH/RB1CC1 coiled-coil complex and unravel the detailed binding mechanism of TAX1BP1 SKICH with RB1CC1. Moreover, we demonstrate that RB1CC1 and NAP1 are competitive in binding to the TAX1BP1 SKICH domain, but the presence of NAP1's FIP200-interacting region (FIR) motif can stabilize the ternary TAX1BP1/NAP1/RB1CC1 complex formation. Finally, we elucidate the molecular mechanism governing the selective interactions of TAX1BP1 with ATG8 family members by solving the structure of GABARAP in complex with the non-canonical LIR (LC3-interacting region) motif of TAX1BP1, which unveils a unique binding mode between LIR and ATG8 family protein. Collectively, our findings provide mechanistic insights into the interactions of TAX1BP1 with RB1CC1 and mammalian ATG8 family proteins and are valuable for further understanding the working mode and function of TAX1BP1 in autophagy.
    Keywords:  GABARAP; RB1CC1; TAX1BP1; autophagy receptor; selective autophagy
    DOI:  https://doi.org/10.1073/pnas.2315550121
  8. Sci Adv. 2024 Mar 08. 10(10): eadn2706
    Proteostatic tuning underpins the evolution of novel multicellular traits.
    Kristopher Montrose, Dung T Lac, Anthony J Burnetti, Kai Tong, G Ozan Bozdag, Mikaela Hukkanen, William C Ratcliff, Juha Saarikangas.
      The evolution of multicellularity paved the way for the origin of complex life on Earth, but little is known about the mechanistic basis of early multicellular evolution. Here, we examine the molecular basis of multicellular adaptation in the multicellularity long-term evolution experiment (MuLTEE). We demonstrate that cellular elongation, a key adaptation underpinning increased biophysical toughness and organismal size, is convergently driven by down-regulation of the chaperone Hsp90. Mechanistically, Hsp90-mediated morphogenesis operates by destabilizing the cyclin-dependent kinase Cdc28, resulting in delayed mitosis and prolonged polarized growth. Reinstatement of Hsp90 or Cdc28 expression resulted in shortened cells that formed smaller groups with reduced multicellular fitness. Together, our results show how ancient protein folding systems can be tuned to drive rapid evolution at a new level of biological individuality by revealing novel developmental phenotypes.
    DOI:  https://doi.org/10.1126/sciadv.adn2706
  9. Life Sci Alliance. 2024 May;pii: e202302370. [Epub ahead of print]7(5):
    Tight regulation of a nuclear HAPSTR1-HUWE1 pathway essential for mammalian life.
    David R Amici, Sammy Alhayek, Austin T Klein, Yi-Zhi Wang, Anika P Wilen, Weimin Song, Pei Zhu, Abhishek Thakkar, McKenzi A King, Adam Wt Steffeck, Milad J Alasady, Clara Peek, Jeffrey N Savas, Marc L Mendillo.
      The recently discovered HAPSTR1 protein broadly oversees cellular stress responses. This function requires HUWE1, a ubiquitin ligase that paradoxically marks HAPSTR1 for degradation, but much about this pathway remains unclear. Here, leveraging multiplexed proteomics, we find that HAPSTR1 enables nuclear localization of HUWE1 with implications for nuclear protein quality control. We show that HAPSTR1 is tightly regulated and identify ubiquitin ligase TRIP12 and deubiquitinase USP7 as upstream regulators titrating HAPSTR1 stability. Finally, we generate conditional Hapstr1 knockout mice, finding that Hapstr1-null mice are perinatal lethal, adult mice depleted of Hapstr1 have reduced fitness, and primary cells explanted from Hapstr1-null animals falter in culture coincident with HUWE1 mislocalization and broadly remodeled signaling. Notably, although HAPSTR1 potently suppresses p53, we find that Hapstr1 is essential for life even in mice lacking p53. Altogether, we identify novel components and functional insights into the conserved HAPSTR1-HUWE1 pathway and demonstrate its requirement for mammalian life.
    DOI:  https://doi.org/10.26508/lsa.202302370
  10. Autophagy. 2024 Mar 05.
    International consensus guidelines for the definition, detection, and interpretation of autophagy-dependent ferroptosis.
    Xin Chen, Andrey S Tsvetkov, Han-Ming Shen, Ciro Isidoro, Nicholas T Ktistakis, Andreas Linkermann, Werner J H Koopman, Hans-Uwe Simon, Lorenzo Galluzzi, Shouqing Luo, Daqian Xu, Wei Gu, Olivier Peulen, Qian Cai, David C Rubinsztein, Jen-Tsan Chi, Donna D Zhang, Changfeng Li, Shinya Toyokuni, Jinbao Liu, Jong-Lyel Roh, Enyong Dai, Gabor Juhasz, Wei Liu, Jianhua Zhang, Minghua Yang, Jiao Liu, Ling-Qiang Zhu, Weiping Zou, Mauro Piacentini, Wen-Xing Ding, Zhenyu Yue, Yangchun Xie, Morten Petersen, David A Gewirtz, Michael A Mandell, Charleen T Chu, Debasish Sinha, Eftekhar Eftekharpour, Boris Zhivotovsky, Sébastien Besteiro, Dmitry I Gabrilovich, Do-Hyung Kim, Valerian E Kagan, Hülya Bayir, Guang-Chao Chen, Scott Ayton, Jan D Lünemann, Masaaki Komatsu, Stefan Krautwald, Ben Loos, Eric H Baehrecke, Jiayi Wang, Jon D Lane, Junichi Sadoshima, Wan Seok Yang, Minghui Gao, Christian Münz, Michael Thumm, Martin Kampmann, Di Yu, Marta M Lipinski, Jace W Jones, Xuejun Jiang, Herbert J Zeh, Rui Kang, Daniel J Klionsky, Guido Kroemer, Daolin Tang.
      Macroautophagy/autophagy is a complex degradation process with a dual role in cell death that is influenced by the cell types that are involved and the stressors they are exposed to. Ferroptosis is an iron-dependent oxidative form of cell death characterized by unrestricted lipid peroxidation in the context of heterogeneous and plastic mechanisms. Recent studies have shed light on the involvement of specific types of autophagy (e.g. ferritinophagy, lipophagy, and clockophagy) in initiating or executing ferroptotic cell death through the selective degradation of anti-injury proteins or organelles. Conversely, other forms of selective autophagy (e.g. reticulophagy and lysophagy) enhance the cellular defense against ferroptotic damage. Dysregulated autophagy-dependent ferroptosis has implications for a diverse range of pathological conditions. This review aims to present an updated definition of autophagy-dependent ferroptosis, discuss influential substrates and receptors, outline experimental methods, and propose guidelines for interpreting the results.
    Keywords:  Cell death; ferritinophagy; iron; lipid peroxidation; lipophagy; lysosome
    DOI:  https://doi.org/10.1080/15548627.2024.2319901
  11. iScience. 2024 Mar 15. 27(3): 109302
    Mapping the substrate landscape of protein phosphatase 2A catalytic subunit PPP2CA.
    Abigail Brewer, Gajanan Sathe, Billie E Pflug, Rosemary G Clarke, Thomas J Macartney, Gopal P Sapkota.
      Protein phosphatase 2A (PP2A) is an essential Ser/Thr phosphatase. The PP2A holoenzyme complex comprises a scaffolding (A), regulatory (B), and catalytic (C) subunit, with PPP2CA being the principal catalytic subunit. The full scope of PP2A substrates in cells remains to be defined. To address this, we employed dTAG proteolysis-targeting chimeras to efficiently and selectively degrade dTAG-PPP2CA in homozygous knock-in HEK293 cells. Unbiased global phospho-proteomics identified 2,204 proteins with significantly increased phosphorylation upon dTAG-PPP2CA degradation, implicating them as potential PPP2CA substrates. A vast majority of these are novel. Bioinformatic analyses revealed involvement of the potential PPP2CA substrates in spliceosome function, cell cycle, RNA transport, and ubiquitin-mediated proteolysis. We identify a pSP/pTP motif as a predominant target for PPP2CA and confirm some of our phospho-proteomic data with immunoblotting. We provide an in-depth atlas of potential PPP2CA substrates and establish targeted degradation as a robust tool to unveil phosphatase substrates in cells.
    Keywords:  Enzymology; Properties of biomolecules; Protein; Proteomics
    DOI:  https://doi.org/10.1016/j.isci.2024.109302
  12. Life Sci Alliance. 2024 May;pii: e202302245. [Epub ahead of print]7(5):
    Oncogenic mutations of KRAS modulate its turnover by the CUL3/LZTR1 E3 ligase complex.
    Andreas Damianou, Zhu Liang, Frederik Lassen, Iolanda Vendrell, George Vere, Svenja Hester, Philip D Charles, Adan Pinto-Fernandez, Alberto Santos, Roman Fischer, Benedikt M Kessler.
      KRAS is a proto-oncogene encoding a small GTPase. Mutations contribute to ∼30% of human solid tumours, including lung adenocarcinoma, pancreatic, and colorectal carcinomas. Most KRAS activating mutations interfere with GTP hydrolysis, essential for its role as a molecular switch, leading to alterations in their molecular environment and oncogenic signalling. However, the precise signalling cascades these mutations affect are poorly understood. Here, APEX2 proximity labelling was used to profile the molecular environment of WT, G12D, G13D, and Q61H-activating KRAS mutants under starvation and stimulation conditions. Through quantitative proteomics, we demonstrate the presence of known KRAS interactors, including ARAF and LZTR1, which are differentially captured by WT and KRAS mutants. Notably, the KRAS mutations G12D, G13D, and Q61H abrogate their association with LZTR1, thereby affecting turnover. Elucidating the implications of LZTR1-mediated regulation of KRAS protein levels in cancer may offer insights into therapeutic strategies targeting KRAS-driven malignancies.
    DOI:  https://doi.org/10.26508/lsa.202302245
  13. Biochem J. 2024 Mar 05. pii: BCJ20220531. [Epub ahead of print]
    Evidence that Xrn1 is in complex with Gcn1, and is required for full levels of eIF2α phosphorylation.
    Renuka Shanmugam, Reuben Anderson, Anja H Schiemann, Evelyn Sattlegger.
      The protein kinase Gcn2 and its effector protein Gcn1 are part of the General Amino Acid Control signalling (GAAC) pathway best known in yeast for its function in maintaining amino acid homeostasis.  Under amino acid limitation, Gcn2 becomes activated, subsequently increasing the levels of phosphorylated eIF2α (eIF2α-P).  This leads to the increased translation of transcriptional regulators, such as Gcn4 in yeast and ATF4 in mammals, and subsequent re-programming of the cell's gene transcription profile, thereby allowing cells to cope with starvation.  Xrn1 is involved in RNA decay, quality control and processing.  We found that Xrn1 co-precipitates Gcn1 and Gcn2, suggesting that these three proteins are in the same complex.  Growth under starvation conditions was dependent on Xrn1 but not on Xrn1-ribosome association, and this correlated with reduced eIF2α-P levels.  Constitutively active Gcn2 leads to a growth defect due to eIF2α-hyperphosphorylation, and we found that this phenotype was independent of Xrn1, suggesting that xrn1 deletion doesn't enhance eIF2α de-phosphorylation.  Our study provides evidence that Xrn1 is required for efficient Gcn2 activation, directly or indirectly.  Thus, we have uncovered a potential new link between RNA metabolism and the GAAC.
    Keywords:  Gcn1; Gcn2; Molecular Biology; Sacharomyces cerevisiae; Xrn1; starvation signaling
    DOI:  https://doi.org/10.1042/BCJ20220531
  14. Nat Cell Biol. 2024 Mar 07.
    Loss of WIPI4 in neurodegeneration causes autophagy-independent ferroptosis.
    Ye Zhu, Motoki Fujimaki, Louisa Snape, Ana Lopez, Angeleen Fleming, David C Rubinsztein.
      β-Propeller protein-associated neurodegeneration (BPAN) is a rare X-linked dominant disease, one of several conditions that manifest with neurodegeneration and brain iron accumulation. Mutations in the WD repeat domain 45 (WDR45) gene encoding WIPI4 lead to loss of function in BPAN but the cellular mechanisms of how these trigger pathology are unclear. The prevailing view in the literature is that BPAN is simply the consequence of autophagy deficiency given that WIPI4 functions in this degradation pathway. However, our data indicate that WIPI4 depletion causes ferroptosis-a type of cell death induced by lipid peroxidation-via an autophagy-independent mechanism, as demonstrated both in cell culture and in zebrafish. WIPI4 depletion increases ATG2A localization at endoplasmic reticulum-mitochondrial contact sites, which enhances phosphatidylserine import into mitochondria. This results in increased mitochondrial synthesis of phosphatidylethanolamine, a major lipid prone to peroxidation, thus enabling ferroptosis. This mechanism has minimal overlap with classical ferroptosis stimuli but provides insights into the causes of neurodegeneration in BPAN and may provide clues for therapeutic strategies.
    DOI:  https://doi.org/10.1038/s41556-024-01373-3
  15. Nat Commun. 2024 Mar 07. 15(1): 1528
    RHBDL4-triggered downregulation of COPII adaptor protein TMED7 suppresses TLR4-mediated inflammatory signaling.
    Julia D Knopf, Susanne S Steigleder, Friederike Korn, Nathalie Kühnle, Marina Badenes, Marina Tauber, Sebastian J Theobald, Jan Rybniker, Colin Adrain, Marius K Lemberg.
      The toll-like receptor 4 (TLR4) is a central regulator of innate immunity that primarily recognizes bacterial lipopolysaccharide cell wall constituents to trigger cytokine secretion. We identify the intramembrane protease RHBDL4 as a negative regulator of TLR4 signaling. We show that RHBDL4 triggers degradation of TLR4's trafficking factor TMED7. This counteracts TLR4 transport to the cell surface. Notably, TLR4 activation mediates transcriptional upregulation of RHBDL4 thereby inducing a negative feedback loop to reduce TLR4 trafficking to the plasma membrane. This secretory cargo tuning mechanism prevents the over-activation of TLR4-dependent signaling in an in vitro Mycobacterium tuberculosis macrophage infection model and consequently alleviates septic shock in a mouse model. A hypomorphic RHBDL4 mutation linked to Kawasaki syndrome, an ill-defined inflammatory disorder in children, further supports the pathophysiological relevance of our findings. In this work, we identify an RHBDL4-mediated axis that acts as a rheostat to prevent over-activation of the TLR4 pathway.
    DOI:  https://doi.org/10.1038/s41467-024-45615-2
  16. iScience. 2024 Mar 15. 27(3): 109235
    Beyond microtubules: The cellular environment at the endoplasmic reticulum attracts proteins to the nucleus, enabling nuclear transport.
    Seok Joo Chae, Dae Wook Kim, Oleg A Igoshin, Seunggyu Lee, Jae Kyoung Kim.
      All proteins are translated in the cytoplasm, yet many, including transcription factors, play vital roles in the nucleus. While previous research has concentrated on molecular motors for the transport of these proteins to the nucleus, recent observations reveal perinuclear accumulation even in the absence of an energy source, hinting at alternative mechanisms. Here, we propose that structural properties of the cellular environment, specifically the endoplasmic reticulum (ER), can promote molecular transport to the perinucleus without requiring additional energy expenditure. Specifically, physical interaction between proteins and the ER impedes their diffusion and leads to their accumulation near the nucleus. This result explains why larger proteins, more frequently interacting with the ER membrane, tend to accumulate at the perinucleus. Interestingly, such diffusion in a heterogeneous environment follows Chapman's law rather than the popular Fick's law. Our findings suggest a novel protein transport mechanism arising solely from characteristics of the intracellular environment.
    Keywords:  Biophysics; Cell biology; Computer simulation
    DOI:  https://doi.org/10.1016/j.isci.2024.109235
  17. Autophagy. 2024 Mar 08. 1-2
    Identification of the YIPF3-YIPF4 heterodimer as a novel Golgiphagy receptor.
    Yuxiang Huang, Daniel J Klionsky.
      Golgiphagy is a selective form of macroautophagy, characterized by the targeted degradation of Golgi compartments through specific receptors. In two recent studies, the YIPF3-YIPF4 heterodimer has been independently identified as the first Golgiphagy receptor within mammalian cells. This heterodimeric complex exhibits a direct affinity for mammalian Atg8-family proteins (ATG8s), thereby facilitating the expansion of phagophores in proximity to Golgi regions. Notably, the interaction between YIPF3-YIPF4 heterodimers and ATG8s undergoes regulatory modulation through phosphorylation. Furthermore, cells lacking either YIPF3 or YIPF4 display defects in Golgiphagy. To elucidate the physiological relevance of these proteins, the necessity of YIPF3-YIPF4 in orchestrating Golgi proteome remodeling was substantiated through experimentation in an in vitro neuronal differentiation model.Abbreviation: ATG: autophagy related; ATG8s: mammalian Atg8-family proteins; LIR, LC3-interacting region.
    Keywords:  Autophagy; LC3-interacting region/LIR; lysosome; proteomics; stress
    DOI:  https://doi.org/10.1080/15548627.2024.2323297
  18. Elife. 2024 Mar 06. pii: e85962. [Epub ahead of print]13
    The ER tether VAPA is required for proper cell motility and anchors ER-PM contact sites to focal adhesions.
    Hugo Siegfried, Georges Farkouh, Rémi Le Borgne, Catherine Pioche-Durieu, Thaïs De Azevedo Laplace, Agathe Verraes, Lucien Daunas, Jean-Marc Verbavatz, Mélina L Heuzé.
      Cell motility processes highly depend on the membrane distribution of Phosphoinositides, giving rise to cytoskeleton reshaping and membrane trafficking events. Membrane contact sites serve as platforms for direct lipid exchange and calcium fluxes between two organelles. Here, we show that VAPA, an ER transmembrane contact site tether, plays a crucial role during cell motility. CaCo2 adenocarcinoma epithelial cells depleted for VAPA exhibit several collective and individual motility defects, disorganized actin cytoskeleton and altered protrusive activity. During migration, VAPA is required for the maintenance of PI(4)P and PI(4,5)P2 levels at the plasma membrane, but not for PI(4)P homeostasis in the Golgi and endosomal compartments. Importantly, we show that VAPA regulates the dynamics of focal adhesions (FA) through its MSP domain, is essential to stabilize and anchor ventral ER-PM contact sites to FA, and mediates microtubule-dependent FA disassembly. To conclude, our results reveal unknown functions for VAPA-mediated membrane contact sites during cell motility and provide a dynamic picture of ER-PM contact sites connection with FA mediated by VAPA.
    Keywords:  E. coli; cell adhesion; cell biology; cell migration; endoplasmic reticulum; human; lipid trafficking; membrane contact sites; phosphoinositides
    DOI:  https://doi.org/10.7554/eLife.85962
  19. Nat Commun. 2024 Mar 08. 15(1): 1908
    Deficiency of ASGR1 promotes liver injury by increasing GP73-mediated hepatic endoplasmic reticulum stress.
    Zhe Zhang, Xiang Kai Leng, Yuan Yuan Zhai, Xiao Zhang, Zhi Wei Sun, Jun Ying Xiao, Jun Feng Lu, Kun Liu, Bo Xia, Qi Gao, Miao Jia, Cheng Qi Xu, Yi Na Jiang, Xiao Gang Zhang, Kai Shan Tao, Jiang Wei Wu.
      Liver injury is a core pathological process in the majority of liver diseases, yet the genetic factors predisposing individuals to its initiation and progression remain poorly understood. Here we show that asialoglycoprotein receptor 1 (ASGR1), a lectin specifically expressed in the liver, is downregulated in patients with liver fibrosis or cirrhosis and male mice with liver injury. ASGR1 deficiency exacerbates while its overexpression mitigates acetaminophen-induced acute and CCl4-induced chronic liver injuries in male mice. Mechanistically, ASGR1 binds to an endoplasmic reticulum stress mediator GP73 and facilitates its lysosomal degradation. ASGR1 depletion increases circulating GP73 levels and promotes the interaction between GP73 and BIP to activate endoplasmic reticulum stress, leading to liver injury. Neutralization of GP73 not only attenuates ASGR1 deficiency-induced liver injuries but also improves survival in mice received a lethal dose of acetaminophen. Collectively, these findings identify ASGR1 as a potential genetic determinant of susceptibility to liver injury and propose it as a therapeutic target for the treatment of liver injury.
    DOI:  https://doi.org/10.1038/s41467-024-46135-9
  20. Nat Commun. 2024 Mar 05. 15(1): 2011
    Riboformer: a deep learning framework for predicting context-dependent translation dynamics.
    Bin Shao, Jiawei Yan, Jing Zhang, Lili Liu, Ye Chen, Allen R Buskirk.
      Translation elongation is essential for maintaining cellular proteostasis, and alterations in the translational landscape are associated with a range of diseases. Ribosome profiling allows detailed measurements of translation at the genome scale. However, it remains unclear how to disentangle biological variations from technical artifacts in these data and identify sequence determinants of translation dysregulation. Here we present Riboformer, a deep learning-based framework for modeling context-dependent changes in translation dynamics. Riboformer leverages the transformer architecture to accurately predict ribosome densities at codon resolution. When trained on an unbiased dataset, Riboformer corrects experimental artifacts in previously unseen datasets, which reveals subtle differences in synonymous codon translation and uncovers a bottleneck in translation elongation. Further, we show that Riboformer can be combined with in silico mutagenesis to identify sequence motifs that contribute to ribosome stalling across various biological contexts, including aging and viral infection. Our tool offers a context-aware and interpretable approach for standardizing ribosome profiling datasets and elucidating the regulatory basis of translation kinetics.
    DOI:  https://doi.org/10.1038/s41467-024-46241-8
  21. Commun Biol. 2024 Mar 02. 7(1): 260
    Structural dynamics of RAF1-HSP90-CDC37 and HSP90 complexes reveal asymmetric client interactions and key structural elements.
    Lorenzo I Finci, Mayukh Chakrabarti, Gulcin Gulten, Joseph Finney, Carissa Grose, Tara Fox, Renbin Yang, Dwight V Nissley, Frank McCormick, Dominic Esposito, Trent E Balius, Dhirendra K Simanshu.
      RAF kinases are integral to the RAS-MAPK signaling pathway, and proper RAF1 folding relies on its interaction with the chaperone HSP90 and the cochaperone CDC37. Understanding the intricate molecular interactions governing RAF1 folding is crucial for comprehending this process. Here, we present a cryo-EM structure of the closed-state RAF1-HSP90-CDC37 complex, where the C-lobe of the RAF1 kinase domain binds to one side of the HSP90 dimer, and an unfolded N-lobe segment of the RAF1 kinase domain threads through the center of the HSP90 dimer. CDC37 binds to the kinase C-lobe, mimicking the N-lobe with its HxNI motif. We also describe structures of HSP90 dimers without RAF1 and CDC37, displaying only N-terminal and middle domains, which we term the semi-open state. Employing 1 μs atomistic simulations, energetic decomposition, and comparative structural analysis, we elucidate the dynamics and interactions within these complexes. Our quantitative analysis reveals that CDC37 bridges the HSP90-RAF1 interaction, RAF1 binds HSP90 asymmetrically, and that HSP90 structural elements engage RAF1's unfolded region. Additionally, N- and C-terminal interactions stabilize HSP90 dimers, and molecular interactions in HSP90 dimers rearrange between the closed and semi-open states. Our findings provide valuable insight into the contributions of HSP90 and CDC37 in mediating client folding.
    DOI:  https://doi.org/10.1038/s42003-024-05959-3
  22. PLoS Genet. 2024 Mar 04. 20(3): e1011169
    Basement membrane diversification relies on two competitive secretory routes defined by Rab10 and Rab8 and modulated by dystrophin and the exocyst complex.
    Cynthia Dennis, Pierre Pouchin, Graziella Richard, Vincent Mirouse.
      The basement membrane (BM) is an essential structural element of tissues, and its diversification participates in organ morphogenesis. However, the traffic routes associated with BM formation and the mechanistic modulations explaining its diversification are still poorly understood. Drosophila melanogaster follicular epithelium relies on a BM composed of oriented BM fibrils and a more homogenous matrix. Here, we determined the specific molecular identity and cell exit sites of BM protein secretory routes. First, we found that Rab10 and Rab8 define two parallel routes for BM protein secretion. When both routes were abolished, BM production was fully blocked; however, genetic interactions revealed that these two routes competed. Rab10 promoted lateral and planar-polarized secretion, whereas Rab8 promoted basal secretion, leading to the formation of BM fibrils and homogenous BM, respectively. We also found that the dystrophin-associated protein complex (DAPC) and Rab10 were both present in a planar-polarized tubular compartment containing BM proteins. DAPC was essential for fibril formation and sufficient to reorient secretion towards the Rab10 route. Moreover, we identified a dual function for the exocyst complex in this context. First, the Exo70 subunit directly interacted with dystrophin to limit its planar polarization. Second, the exocyst complex was also required for the Rab8 route. Altogether, these results highlight important mechanistic aspects of BM protein secretion and illustrate how BM diversity can emerge from the spatial control of distinct traffic routes.
    DOI:  https://doi.org/10.1371/journal.pgen.1011169
  23. Science. 2024 Mar 07. eadl2528
    Generalized biomolecular modeling and design with RoseTTAFold All-Atom.
    Rohith Krishna, Jue Wang, Woody Ahern, Pascal Sturmfels, Preetham Venkatesh, Indrek Kalvet, Gyu Rie Lee, Felix S Morey-Burrows, Ivan Anishchenko, Ian R Humphreys, Ryan McHugh, Dionne Vafeados, Xinting Li, George A Sutherland, Andrew Hitchcock, C Neil Hunter, Alex Kang, Evans Brackenbrough, Asim K Bera, Minkyung Baek, Frank DiMaio, David Baker.
      Deep learning methods have revolutionized protein structure prediction and design but are currently limited to protein-only systems. We describe RoseTTAFold All-Atom (RFAA) which combines a residue-based representation of amino acids and DNA bases with an atomic representation of all other groups to model assemblies containing proteins, nucleic acids, small molecules, metals, and covalent modifications given their sequences and chemical structures. By fine tuning on denoising tasks we obtain RFdiffusionAA, which builds protein structures around small molecules. Starting from random distributions of amino acid residues surrounding target small molecules, we design and experimentally validate, through crystallography and binding measurements, proteins that bind the cardiac disease therapeutic digoxigenin, the enzymatic cofactor heme, and the light harvesting molecule bilin.
    DOI:  https://doi.org/10.1126/science.adl2528
  24. PLoS Biol. 2024 Mar;22(3): e3002537
    Pharmacological induction of autophagy reduces inflammation in macrophages by degrading immunoproteasome subunits.
    Jiao Zhou, Chunxia Li, Meng Lu, Gaoyue Jiang, Shanze Chen, Huihui Li, Kefeng Lu.
      Defective autophagy is linked to proinflammatory diseases. However, the mechanisms by which autophagy limits inflammation remain elusive. Here, we found that the pan-FGFR inhibitor LY2874455 efficiently activated autophagy and suppressed expression of proinflammatory factors in macrophages stimulated by lipopolysaccharide (LPS). Multiplex proteomic profiling identified the immunoproteasome, which is a specific isoform of the 20s constitutive proteasome, as a substrate that is degraded by selective autophagy. SQSTM1/p62 was found to be a selective autophagy-related receptor that mediated this degradation. Autophagy deficiency or p62 knockdown blocked the effects of LY2874455, leading to the accumulation of immunoproteasomes and increases in inflammatory reactions. Expression of proinflammatory factors in autophagy-deficient macrophages could be reversed by immunoproteasome inhibitors, confirming the pivotal role of immunoproteasome turnover in the autophagy-mediated suppression on the expression of proinflammatory factors. In mice, LY2874455 protected against LPS-induced acute lung injury and dextran sulfate sodium (DSS)-induced colitis and caused low levels of proinflammatory cytokines and immunoproteasomes. These findings suggested that selective autophagy of the immunoproteasome was a key regulator of signaling via the innate immune system.
    DOI:  https://doi.org/10.1371/journal.pbio.3002537
  25. Heliyon. 2024 Mar 15. 10(5): e25937
    Molecular mechanism of ATF6 in unfolded protein response and its role in disease.
    Yingying Lei, Hong Yu, Shaoxue Ding, Hui Liu, Chunyan Liu, Rong Fu.
      Activating transcription factor 6 (ATF6), an important signaling molecule in unfolded protein response (UPR), plays a role in the pathogenesis of several diseases, including diseases such as congenital retinal disease, liver fibrosis and ankylosing spondylitis. After endoplasmic reticulum stress (ERS), ATF6 is activated after separation from binding immunoglobulin protein (GRP78/BiP) in the endoplasmic reticulum (ER) and transported to the Golgi apparatus to be hydrolyzed by site 1 and site 2 proteases into ATF6 fragments, which localize to the nucleus and regulate the transcription and expression of ERS-related genes. In these diseases, ERS leads to the activation of UPR, which ultimately lead to the occurrence and development of diseases by regulating the physiological state of cells through the ATF6 signaling pathway. Here, we discuss the evidence for the pathogenic importance of ATF6 signaling in different diseases and discuss preclinical results.
    Keywords:  Activating transcription factor 6 (ATF6); Endoplasmic reticulum stress (ERS); Proteostasis; Unfolded protein response (UPR)
    DOI:  https://doi.org/10.1016/j.heliyon.2024.e25937
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