cometadata/datacite-titles-descriptions-related-identifiers huggingface dataset:
DOI: 10.6084/m9.figshare.29924822.v1
title: Inhibition of UBE2N promotes the clearance of mutant HTT (huntingtin) in HD knock-in mice

description: Accumulation of misfolded proteins leads to many neurodegenerative diseases that can be treated by lowering or removing mutant proteins. Huntington disease (HD) is characterized by the accumulation of ubiquitinated mutant HTT (huntingtin) in the central nervous system. Ubiquitination of the misfolded proteins, a common feature of the neurodegenerative diseases, is mediated by the different lysine residues on ubiquitin. We previously discovered that the age-dependent increase of UBE2N (ubiquitin conjugating enzyme E2 N) exacerbated the accumulation of misfolded HTT and amyloid proteins, accompanied by the elevation of K63 ubiquitination. Pharmacological inhibition of UBE2N could ameliorate the amyloid deposition. However, the effect of UBE2N suppression on HTT aggregate clearance has remained unknown. In the current work, we demonstrate that selectively suppressing UBE2N, with antisense oligonucleotides or small-molecular inhibitors, increased removal of HTT aggregates by proteasome degradation in the striatum of HD knock-in mice. We also identified two novel ubiquitin specific peptidases, USP29 and USP49, that participated in the clearance of HTT aggregates, via accelerating K48-mediated ubiquitin-proteasome function. Our findings provide a potential pharmacological approach to treat neurodegeneration caused by mutant HTT. <b>Abbreviation</b>: AD: Alzheimer disease; ALP: autophagy-lysosomal pathway; AMC: 7-Amino-4-Methylcoumarin; ASO: antisense oligonucleotide; Aβ: amyloid β; BafA1: bafilomycin A<sub>1</sub>; DEG: differentially expressed gene; DMSO: dimethyl sulfoxide; E2: ubiquitin-conjugating; E3: ubiquitin-ligating; EGFP: enhanced green fluorescent protein; HD: Huntington disease; HTT: huntingtin; KI: knock-in; SDS: sodium dodecyl sulfate; UPS: ubiquitin-proteasome system; UBE2N: ubiquitin conjugating enzyme E2 N; USP: ubiquitin-specific peptidase.
DOI: 10.6084/m9.figshare.30382802
title: Lysosomal proteomics reveals mechanisms of neuronal APOE4-associated lysosomal dysfunction

description: APOE4 is the primary risk factor for Alzheimer disease (AD). Early AD pathological events first affect the neuronal endolysosomal system, which in turn causes neuronal protein aggregation and cell death. Despite the crucial influence of lysosomes upon AD pathophysiology, and that APOE4 localizes to lysosomes, the influence of APOE4 on lysosomal function remains unexplored. We find that expression of APOE4 in neuronal cell lines results in lysosomal alkalinization and impaired lysosomal function. To identify driving factors for these defects, we performed quantitative lysosomal proteome profiling. This revealed that APOE4 expression results in differential regulation of numerous lysosomal proteins, correlating with APOE allele status and disease severity in AD brains. In particular, APOE4 expression results in the depletion of lysosomal LGALS3BP and the accumulation of lysosomal TMED5. We additionally validated that these lysosomal protein changes can be targeted to modulate lysosomal function. Taken together, this work thereby reveals that APOE4 causes widespread lysosomal defects through remodeling the lysosomal proteome, with the lysosomal TMED5 accumulation and LGALS3BP depletion manifesting as lysosomal alkalinization in APOE4 neurons. <b>Abbreviation</b>: Aβ-β-amyloid; AD-Alzheimer disease; APMS-affinity purification mass spectrometry; APOE-apolipoprotein E; APP-amyloid beta precursor protein; BafA1-bafilomycin A1; CMA-chaperone-mediated autophagy; COP-coatomer; ER-endoplasmic reticulum; ERAD-endoplasmic reticulum-associated degradation; ERGIC-endoplasmic reticulum-Golgi intermediate compartment; FCCP-carbonyl cyanide p-trifluoromethoxyphenylhydrazone; GABA-gamma-aminobutyric acid; GFP-green fluorescent protein; GPI-glycerophosphoinositol; GPN-glycyl-L-phenylalanine 2-naphthylamide; HA-hemagglutinin protein tag; iPSC-induced pluripotent stem cells; LAMP1-lysosomal associated membrane protein 1; LGALS3BP-galectin 3 binding protein; LysoIP-lysosomal immunoprecipitation; MAPT-microtubule associated protein tau; MR-CTSB-magic red cathepsin B; MS-mass spectrometry; NFT-neurofibrillary tangles; PCA-principal component analysis; PLA-proximity ligation assay; PSEN1-presenilin 1; PSEN2-presenilin 2; p-MAPT/Tau-phospho-MAPT/tau; ROS-reactive oxygen species; RT-qPCR-reverse transcriptase quantitative polymerase chain reaction; SEM-standard error of the mean; siRNA-short interfering RNA; SRCR-scavenger receptor cysteine-rich domain; TMED-transmembrane p24 trafficking protein; TMEM192-transmembrane protein 192
DOI: 10.6084/m9.figshare.30382802.v1
title: Lysosomal proteomics reveals mechanisms of neuronal APOE4-associated lysosomal dysfunction

description: APOE4 is the primary risk factor for Alzheimer disease (AD). Early AD pathological events first affect the neuronal endolysosomal system, which in turn causes neuronal protein aggregation and cell death. Despite the crucial influence of lysosomes upon AD pathophysiology, and that APOE4 localizes to lysosomes, the influence of APOE4 on lysosomal function remains unexplored. We find that expression of APOE4 in neuronal cell lines results in lysosomal alkalinization and impaired lysosomal function. To identify driving factors for these defects, we performed quantitative lysosomal proteome profiling. This revealed that APOE4 expression results in differential regulation of numerous lysosomal proteins, correlating with APOE allele status and disease severity in AD brains. In particular, APOE4 expression results in the depletion of lysosomal LGALS3BP and the accumulation of lysosomal TMED5. We additionally validated that these lysosomal protein changes can be targeted to modulate lysosomal function. Taken together, this work thereby reveals that APOE4 causes widespread lysosomal defects through remodeling the lysosomal proteome, with the lysosomal TMED5 accumulation and LGALS3BP depletion manifesting as lysosomal alkalinization in APOE4 neurons. <b>Abbreviation</b>: Aβ-β-amyloid; AD-Alzheimer disease; APMS-affinity purification mass spectrometry; APOE-apolipoprotein E; APP-amyloid beta precursor protein; BafA1-bafilomycin A1; CMA-chaperone-mediated autophagy; COP-coatomer; ER-endoplasmic reticulum; ERAD-endoplasmic reticulum-associated degradation; ERGIC-endoplasmic reticulum-Golgi intermediate compartment; FCCP-carbonyl cyanide p-trifluoromethoxyphenylhydrazone; GABA-gamma-aminobutyric acid; GFP-green fluorescent protein; GPI-glycerophosphoinositol; GPN-glycyl-L-phenylalanine 2-naphthylamide; HA-hemagglutinin protein tag; iPSC-induced pluripotent stem cells; LAMP1-lysosomal associated membrane protein 1; LGALS3BP-galectin 3 binding protein; LysoIP-lysosomal immunoprecipitation; MAPT-microtubule associated protein tau; MR-CTSB-magic red cathepsin B; MS-mass spectrometry; NFT-neurofibrillary tangles; PCA-principal component analysis; PLA-proximity ligation assay; PSEN1-presenilin 1; PSEN2-presenilin 2; p-MAPT/Tau-phospho-MAPT/tau; ROS-reactive oxygen species; RT-qPCR-reverse transcriptase quantitative polymerase chain reaction; SEM-standard error of the mean; siRNA-short interfering RNA; SRCR-scavenger receptor cysteine-rich domain; TMED-transmembrane p24 trafficking protein; TMEM192-transmembrane protein 192
DOI: 10.6084/m9.figshare.29381766
title: Inhibition of lysosomal LAMTOR1 increases autophagy by suppressing the MTORC1 pathway to ameliorate lipid accumulations in MAFLD

description: Metabolic dysfunction-associated fatty liver disease (MAFLD) is a serious metabolic disorder characterized by fat accumulation in the liver, which can trigger liver inflammation and fibrosis, potentially leading to cirrhosis or liver cancer. Despite many studies, effective treatments for MAFLD remain elusive due to its complex etiology. In this study, we have focused on the discovery of therapeutic agents and molecular targets for MAFLD treatment. We demonstrated that the natural compound acacetin (ACA) alleviates MAFLD by regulating macroautophagy/autophagy in a CDAHFD mouse model of rapidly induced steatohepatitis. In addition, ACA inhibits lipid accumulation in 3T3-L1 adipocytes through autophagy induction. To identify the target responsible for the autophagy activity induced by ACA, we performed drug affinity responsive target stability (DARTS) combined with LC-MS/MS proteomic analysis. This led to the identification of LAMTOR1 (late endosomal/lysosomal adaptor, MAPK and MTOR activator 1), a lysosomal membrane adaptor protein. We found that binding of ACA to LAMTOR1 induces its release from the LAMTOR complex, leading to inhibition of MTOR (mechanistic target of rapamycin kinase) complex 1 (MTORC1), thereby increasing autophagy. This process helps ameliorate metabolic disorders by modulating the MTORC1-AMPK axis. Genetic knockdown of LAMTOR1 phenocopies the effects of ACA treatment, further supporting the role of LAMTOR1 as a target of ACA. These findings suggest LAMTOR1 plays a crucial role in ACA’s therapeutic effects on MAFLD. In summary, our study identifies LAMTOR1 as a key protein target of ACA, revealing a potential therapeutic avenue for MAFLD by modulating autophagy via the LAMTOR1-MTORC1-AMPK signaling pathway. <b>Abbreviations:</b> ACA: acacetin; ADGRE1/EMR1/F4/80: adhesion G protein-coupled receptor E1; AMPK: AMP-activated protein kinase; CDAHFD: choline-deficient amino acid-defined, high-fat diet; CETSA: cellular thermal shift assay; CQ: chloroquine; DARTS: drug affinity responsive target stability; DQ-BSA: dye quenched-bovine serum albumin; GOT1/AST: glutamic-oxaloacetic transaminase 1; GPT/ALT: glutamic-pyruvic transaminase; LAMP2: lysosomal associated membrane protein 2; LAMTOR1: late endosomal/lysosomal adaptor, MAPK and MTOR activator 1; LC-MS/MS: liquid chromatography-tandem mass spectrometry; MAFLD: metabolic dysfunction-associated fatty liver disease; MAP1LC3B/LC3: microtubule associated protein 1 light chain 3 beta; MASH: metabolic dysfunction-associated steatohepatitis; mRFP-GFP-MAP1LC3B: tandem fluorescent-tagged MAP1LC3B; MTORC1: mechanistic target of rapamycin complex 1; PA: palmitic acid; PRKAA: protein kinase AMP-activated catalytic subunit alpha; PLA: proximity ligation assay; Rapa: rapamycin; RPS6KB1/p70S6K: ribosomal protein S6 kinase B1; RRAG: Ras-related GTP-binding; SQSTM1: sequestosome 1; TFEB: transcription factor EB; VMP1: vacuole membrane protein 1.
DOI: 10.6084/m9.figshare.29381766.v1
title: Inhibition of lysosomal LAMTOR1 increases autophagy by suppressing the MTORC1 pathway to ameliorate lipid accumulations in MAFLD

description: Metabolic dysfunction-associated fatty liver disease (MAFLD) is a serious metabolic disorder characterized by fat accumulation in the liver, which can trigger liver inflammation and fibrosis, potentially leading to cirrhosis or liver cancer. Despite many studies, effective treatments for MAFLD remain elusive due to its complex etiology. In this study, we have focused on the discovery of therapeutic agents and molecular targets for MAFLD treatment. We demonstrated that the natural compound acacetin (ACA) alleviates MAFLD by regulating macroautophagy/autophagy in a CDAHFD mouse model of rapidly induced steatohepatitis. In addition, ACA inhibits lipid accumulation in 3T3-L1 adipocytes through autophagy induction. To identify the target responsible for the autophagy activity induced by ACA, we performed drug affinity responsive target stability (DARTS) combined with LC-MS/MS proteomic analysis. This led to the identification of LAMTOR1 (late endosomal/lysosomal adaptor, MAPK and MTOR activator 1), a lysosomal membrane adaptor protein. We found that binding of ACA to LAMTOR1 induces its release from the LAMTOR complex, leading to inhibition of MTOR (mechanistic target of rapamycin kinase) complex 1 (MTORC1), thereby increasing autophagy. This process helps ameliorate metabolic disorders by modulating the MTORC1-AMPK axis. Genetic knockdown of LAMTOR1 phenocopies the effects of ACA treatment, further supporting the role of LAMTOR1 as a target of ACA. These findings suggest LAMTOR1 plays a crucial role in ACA’s therapeutic effects on MAFLD. In summary, our study identifies LAMTOR1 as a key protein target of ACA, revealing a potential therapeutic avenue for MAFLD by modulating autophagy via the LAMTOR1-MTORC1-AMPK signaling pathway. <b>Abbreviations:</b> ACA: acacetin; ADGRE1/EMR1/F4/80: adhesion G protein-coupled receptor E1; AMPK: AMP-activated protein kinase; CDAHFD: choline-deficient amino acid-defined, high-fat diet; CETSA: cellular thermal shift assay; CQ: chloroquine; DARTS: drug affinity responsive target stability; DQ-BSA: dye quenched-bovine serum albumin; GOT1/AST: glutamic-oxaloacetic transaminase 1; GPT/ALT: glutamic-pyruvic transaminase; LAMP2: lysosomal associated membrane protein 2; LAMTOR1: late endosomal/lysosomal adaptor, MAPK and MTOR activator 1; LC-MS/MS: liquid chromatography-tandem mass spectrometry; MAFLD: metabolic dysfunction-associated fatty liver disease; MAP1LC3B/LC3: microtubule associated protein 1 light chain 3 beta; MASH: metabolic dysfunction-associated steatohepatitis; mRFP-GFP-MAP1LC3B: tandem fluorescent-tagged MAP1LC3B; MTORC1: mechanistic target of rapamycin complex 1; PA: palmitic acid; PRKAA: protein kinase AMP-activated catalytic subunit alpha; PLA: proximity ligation assay; Rapa: rapamycin; RPS6KB1/p70S6K: ribosomal protein S6 kinase B1; RRAG: Ras-related GTP-binding; SQSTM1: sequestosome 1; TFEB: transcription factor EB; VMP1: vacuole membrane protein 1.
DOI: 10.6084/m9.figshare.29889787
title: A novel mitochondrial regulon for ferroptosis during fungal pathogenesis

description: Ferroptosis remains an underexamined iron- and lipid peroxides-driven cell death modality despite its importance to several human and plant diseases and to immunity thereof. Here, we utilized chemical cell biology, molecular genetics and biochemical analyses to gain insights into how the fungal pathogen <i>Magnaporthe oryzae</i> undergoes ferroptosis strictly in the spore cells to successfully transit to infectious development. We reveal a complex functional interdependency and crosstalk between intrinsic ferroptosis and autophagy-mediated mitochondrial degradation. Mechanistically, the requirement of mitophagy for ferroptotic cell death was attributed to its ability to maintain a pool of metabolically active mitochondria. Pharmacological disruption of the electron transport chain or membrane potential led to complete inhibition of ferroptosis, thus simulating the loss of mitophagy phenotypes. Conversely, increased mitochondrial membrane potential in a mitophagy-defective mutant alleviated the ferroptosis defects therein. Graded inhibition of mitochondrial coenzyme Q biosynthesis with or without ferroptosis inhibitor liproxstatin-1 distinguished its antioxidant function in such regulated cell death. Membrane potential-dependent regulation of ATP synthesis and iron homeostasis, as well as dynamics of tricarboxylic acid cycle enzyme AcoA (aconitase A) in the presence or absence of mitophagy, mitochondrial poisoning or iron chelation further linked mitochondrial metabolism to ferroptosis. Last, we present an important bioenergetics- and redox-based mitochondrial regulon essential for intrinsic ferroptosis and its precise role in fungal pathogenesis leading up to the establishment of the devastating rice blast disease. <b>Abbreviation</b>: 4-CBA: 4 chlorobenzoic acid; AcoA: aconitase A; Atg24: autophagy related 24; CoQ: coenzyme Q; CPX: ciclopirox olamine; ETC: electron transport chain; GSH: glutathione; Gpx4: glutathione peroxidase 4; HPI: hours post inoculation; MMP: mitochondrial membrane potential; MitoQ: Mitoquinone; ROS: reactive oxygen species; TCA: tricarboxylic acid
DOI: 10.6084/m9.figshare.29889787.v2
title: A novel mitochondrial regulon for ferroptosis during fungal pathogenesis

description: Ferroptosis remains an underexamined iron- and lipid peroxides-driven cell death modality despite its importance to several human and plant diseases and to immunity thereof. Here, we utilized chemical cell biology, molecular genetics and biochemical analyses to gain insights into how the fungal pathogen <i>Magnaporthe oryzae</i> undergoes ferroptosis strictly in the spore cells to successfully transit to infectious development. We reveal a complex functional interdependency and crosstalk between intrinsic ferroptosis and autophagy-mediated mitochondrial degradation. Mechanistically, the requirement of mitophagy for ferroptotic cell death was attributed to its ability to maintain a pool of metabolically active mitochondria. Pharmacological disruption of the electron transport chain or membrane potential led to complete inhibition of ferroptosis, thus simulating the loss of mitophagy phenotypes. Conversely, increased mitochondrial membrane potential in a mitophagy-defective mutant alleviated the ferroptosis defects therein. Graded inhibition of mitochondrial coenzyme Q biosynthesis with or without ferroptosis inhibitor liproxstatin-1 distinguished its antioxidant function in such regulated cell death. Membrane potential-dependent regulation of ATP synthesis and iron homeostasis, as well as dynamics of tricarboxylic acid cycle enzyme AcoA (aconitase A) in the presence or absence of mitophagy, mitochondrial poisoning or iron chelation further linked mitochondrial metabolism to ferroptosis. Last, we present an important bioenergetics- and redox-based mitochondrial regulon essential for intrinsic ferroptosis and its precise role in fungal pathogenesis leading up to the establishment of the devastating rice blast disease. <b>Abbreviation</b>: 4-CBA: 4 chlorobenzoic acid; AcoA: aconitase A; Atg24: autophagy related 24; CoQ: coenzyme Q; CPX: ciclopirox olamine; ETC: electron transport chain; GSH: glutathione; Gpx4: glutathione peroxidase 4; HPI: hours post inoculation; MMP: mitochondrial membrane potential; MitoQ: Mitoquinone; ROS: reactive oxygen species; TCA: tricarboxylic acid
DOI: 10.6084/m9.figshare.29889787.v1
title: A novel mitochondrial regulon for ferroptosis during fungal pathogenesis

description: Ferroptosis remains an underexamined iron- and lipid peroxides-driven cell death modality despite its importance to several human and plant diseases and to immunity thereof. Here, we utilized chemical cell biology, molecular genetics and biochemical analyses to gain insights into how the fungal pathogen <i>Magnaporthe oryzae</i> undergoes ferroptosis strictly in the spore cells to successfully transit to infectious development. We reveal a complex functional interdependency and crosstalk between intrinsic ferroptosis and autophagy-mediated mitochondrial degradation. Mechanistically, the requirement of mitophagy for ferroptotic cell death was attributed to its ability to maintain a pool of metabolically active mitochondria. Pharmacological disruption of the electron transport chain or membrane potential led to complete inhibition of ferroptosis, thus simulating the loss of mitophagy phenotypes. Conversely, increased mitochondrial membrane potential in a mitophagy-defective mutant alleviated the ferroptosis defects therein. Graded inhibition of mitochondrial coenzyme Q biosynthesis with or without ferroptosis inhibitor liproxstatin-1 distinguished its antioxidant function in such regulated cell death. Membrane potential-dependent regulation of ATP synthesis and iron homeostasis, as well as dynamics of tricarboxylic acid cycle enzyme AcoA (aconitase A) in the presence or absence of mitophagy, mitochondrial poisoning or iron chelation further linked mitochondrial metabolism to ferroptosis. Last, we present an important bioenergetics- and redox-based mitochondrial regulon essential for intrinsic ferroptosis and its precise role in fungal pathogenesis leading up to the establishment of the devastating rice blast disease.
DOI: 10.6084/m9.figshare.29413487
title: Excessive autophagic degradation of MYLK3 causes sunitinib-induced cardiotoxicity

description: Sunitinib is a receptor tyrosine kinase inhibitor used for the treatment of renal cell carcinoma and imatinib-resistant gastrointestinal stromal tumors. Clinical data have shown that patients receiving sunitinib develop reduced cardiac function, arrhythmia and heart failure, thereby largely limiting its clinical use. However, the molecular mechanisms underlying sunitinib-induced arrhythmogenesis remain unclear. Here, utilizing the human induced pluripotent stem cell-derived cardiomyocyte (iPSC-CM) model, we found that sunitinib caused a variety of deleterious phenotypes, including cardiomyocyte death, sarcomeric disorganization, irregular Ca<sup>2+</sup> transients, impaired ATP2A2a/SERCA2a (ATPase sarcoplasmic/endoplasmic reticulum Ca<sup>2+</sup> transporting 2a) activity, arrhythmia, and excessive macroautophagy/autophagy. Mechanistically, SQSTM1/p62 (sequestosome 1) interacts with MYLK3 (myosin light chain kinase 3) and drives excessive autophagic degradation of MYLK3 in sunitinib-treated iPSC-CMs. Downregulation of MYLK3 suppresses the phosphorylation of CAMK2/CAMKII (calcium/calmodulin dependent protein kinase II), thereby reducing the phosphorylation level of its downstream substrate PLN (phospholamban), leading to impaired ATP2A2a/SERCA2a activity and subsequent Ca<sup>2+</sup> dyshomeostasis and arrhythmia. Moreover, pharmacological intervention of the cardiac myosin activator omecamtiv mecarbil (OM) or overexpression of MYLK3 significantly restored the expression of MYLK3 and reversed pathogenic phenotypes in sunitinib-treated iPSC-CMs. Nanoparticle delivery of OM effectively prevented sunitinib-induced cardiac dysfunction in mice. Our findings suggest that sunitinib-induced MYLK3 degradation causes the inhibition of the CAMK2-PLN-ATP2A2a signaling pathway and leads to sunitinib-induced arrhythmogenesis, and that MYLK3 can act as a novel cardioprotective target for sunitinib-induced cardiotoxicity. <b>Abbreviation</b>: ACTN:actinin alpha;APD:action potential duration; ATG:autophagy related;ATP2A2a/SERCA2a:ATPase sarcoplasmic/endoplasmicreticulum Ca2+ transporting 2a;BafA1:bafilomycin A<sub>1</sub>;Caff: caffine; CAMK2/CAMKII:calcium/calmodulin dependent protein kinase II;CASP3:caspase 3;CQ, chloroquine;DADs:delayed afterdepolarizations; EAD:early afterdepolarization; ECG: electrocardiogram; EF: ejectionfraction; FS: fractional shortening; iPSC:inducedpluripotent stem cell;iPSC-CM: inducedpluripotent stem-cell-derived cardiomyocyte;ISO: isoprenaline; LVIDs: left ventricular end systolic diameter;LVIDd: left ventricular end diastolic diameter;MAP1LC3/LC3:microtubuleassociatedprotein 1 light chain 3;MYL2v/MLC2v:myosin light chain 2 v;MYLK3:myosin light chain kinase 3;OE: overexpression; OM:omecamtiv mecarbil; PLN: phospholamban;SIC:sunitinib-induced cardiotoxicity; SR:sarcoplasmic reticulum; TUNEL:TdT-mediated dUTP nick end labeling.