Autophagy最新文献

Disruption of mitochondrial function is observed in multiple drug-induced liver injuries (DILIs), a significant global health threat. However, how the mitochondrial dysfunction occurs and whether maintain mitochondrial homeostasis is beneficial for DILIs remains unclear. Here, we show that defective mitophagy by OPTN (optineurin) ablation causes disrupted mitochondrial homeostasis and aggravates hepatocytes necrosis in DILIs, while OPTN overexpression protects against DILI depending on its mitophagic function. Notably, mass spectrometry analysis identifies a new mitochondrial substrate, GCDH (glutaryl-CoA dehydrogenase), which can be selectively recruited by OPTN for mitophagic degradation, and a new cofactor, VCP (valosin containing protein) that interacts with OPTN to stabilize BECN1 during phagophore assembly, thus boosting OPTN-mediated mitophagy initiation to clear damaged mitochondria and preserve mitochondrial homeostasis in DILIs. Then, the accumulation of OPTN in different DILIs is further validated with a protective effect, and pyridoxine is screened and established to alleviate DILIs by inducing OPTN-mediated mitophagy. Collectively, our findings uncover a dual role of OPTN in mitophagy initiation and implicate the preservation of mitochondrial homeostasis via inducing OPTN-mediated mitophagy as a potential therapeutic approach for DILIs.Abbreviation: AILI: acetaminophen-induced liver injury; ALS: amyotrophic lateral sclerosis; APAP: acetaminophen; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CHX: cycloheximide; Co-IP: co-immunoprecipitation; DILI: drug-induced liver injury; FL: full length; GCDH: glutaryl-CoA dehydrogenase; GOT1/AST: glutamic-oxaloacetic transaminase 1; GO: gene ontology; GSEA: gene set enrichment analysis; GPT/ALT: glutamic - pyruvic transaminase; INH: isoniazid; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MMP: mitochondrial membrane potential; MST: microscale thermophoresis; MT-CO2/COX-II: mitochondrially encoded cytochrome c oxidase II; OPTN: optineurin; PINK1: PTEN induced kinase 1; PRKN: parkin RBR E3 ubiquitin protein ligase; TIMM23: translocase of inner mitochondrial membrane 23; TOMM20: translocase of outer mitochondrial membrane 20; TSN: toosendanin; VCP: valosin containing protein, WIPI2: WD repeat domain, phosphoinositide interacting 2.

Aging is often accompanied by a decline in proteostasis, manifested as an increased propensity for misfolded protein aggregates, which are prevented by protein quality control systems, such as the ubiquitin-proteasome system (UPS) and macroautophagy/autophagy. Although the role of the UPS and autophagy in slowing age-induced proteostasis decline has been elucidated, limited information is available on how these pathways can be activated in a collaborative manner to delay proteostasis-associated aging. Here, we show that activation of the UPS via the pharmacological inhibition of USP14 (ubiquitin specific peptidase 14) using IU1 improves proteostasis and autophagy decline caused by aging or proteostatic stress in Drosophila and human cells. Treatment with IU1 not only alleviated the aggregation of polyubiquitinated proteins in aging Drosophila flight muscles but also extended the fly lifespan with enhanced locomotive activity via simultaneous activation of the UPS and autophagy. Interestingly, the effect of this drug disappeared when proteasomal activity was inhibited, but was evident upon proteostasis disruption by foxo mutation. Overall, our findings shed light on potential strategies to efficiently ameliorate age-associated pathologies associated with perturbed proteostasis.Abbreviations: AAAs: amino acid analogs; foxo: forkhead box, sub-group O; IFMs: indirect flight muscles; UPS: ubiquitin-proteasome system; USP14: ubiquitin specific peptidase 14.

Macroautophagy/autophagy degrades and recycles cellular constituents via the lysosome to maintain cellular homeostasis. Our study identified the endoplasmic reticulum (ER)-resident SIGMAR1 (sigma non-opioid intracellular receptor 1) as a critical regulator of the biosynthesis of Atg8-family proteins that leads to the lipidation that is essential during autophagosome formation. We demonstrate that SIGMAR1 stabilizes MAP1LC3B/LC3B and GABARAP mRNAs, promoting their localized translation proximal to the ER for efficient lipidation. Using single-molecule fluorescence in situ hybridization/smFISH and co-immunoprecipitation, we found that SIGMAR1 directly binds to a conserved region in the 3' UTR of LC3B mRNA, facilitating its translation, efficient lipidation, and proper integration into the phagophore membrane. Cells lacking SIGMAR1 show reduced levels of many Atg8-family proteins and impaired autophagic flux. Our model suggests that SIGMAR1-mediated localized translation of Atg8-family proteins at the ER promotes efficient autophagosome formation, in contrast to recruiting preexisting cytosolic Atg8-family proteins to the lipidation machinery. Elucidating the role of SIGMAR1 in autophagy may provide better therapeutic strategies to prevent or treat autophagy-dependent neurodegenerative diseases, particularly given the highly druggable nature of SIGMAR1.

The plasma concentration of the macroautophagy/autophagy inhibitor DBI/ACBP (diazepam binding inhibitor, acyl-CoA binding protein) increases with aging and body mass index (BMI). Both advanced age and obesity are among the most important risk factors for the development of cancer. We observed that patients with cancer predisposition syndromes due to mutations in BRCA1, BRCA2 and TP53 exhibit abnormally high plasma DBI/ACBP levels. Additionally, patients without known cancer predisposition syndromes also manifest higher DBI/ACBP levels before imminent cancer diagnosis (within 0-3 years) as compared to age and BMI-matched controls who remain cancer-free. Thus, supranormal plasma DBI/ACBP constitutes a risk factor for later cancer development. Mouse experimentation revealed that genetic or antibody-mediated DBI/ACBP inhibition can delay the development or progression of cancers. In the context of chemoimmunotherapy, DBI/ACBP neutralization enhances tumor infiltration by non-exhausted effector T cells but reduces infiltration by regulatory T cells. This resulted in better cancer control in models of breast cancer, non-small cell lung cancer and sarcoma. We conclude that DBI/ACBP constitutes an actionable autophagy checkpoint for improving cancer immunosurveillance. Abbreviation: BMI, body mass index; CTL, cytotoxic T lymphocyte; DBI, diazepam binding inhibitor, acyl-CoA binding protein; mAb, monoclonal antibody; NSCLC, non-small cell lung cancer; PDCD1/PD-1, programmed cell death 1; scRNA-seq, single-cell RNA sequencing; T_reg, regulatory T cell.

Spautin-1 is a well-known macroautophagy/autophagy inhibitor via suppressing the deubiquitinases USP10 and USP13 and promoting the degradation of the PIK3C3/VPS34-BECN1 complex, while its effect on selective autophagy remains poorly understood. Mitophagy is a selective form of autophagy for removal of damaged and superfluous mitochondria via the autophagy-lysosome pathway. Here, we report a surprising discovery that, while spautin-1 remains as an effective autophagy inhibitor, it promotes PINK1-PRKN-dependent mitophagy induced by mitochondrial damage agents. Mechanistically, spautin-1 facilitates the stabilization and activation of the full-length PINK1 at the outer mitochondrial membrane (OMM) via binding to components of the TOMM complex (TOMM70 and TOMM20), leading to the disruption of the mitochondrial import of PINK1 and prevention of PARL-mediated PINK1 cleavage. Moreover, spautin-1 induces neuronal mitophagy in Caenorhabditis elegans (C. elegans) in a PINK-1-PDR-1-dependent manner. Functionally, spautin-1 is capable of improving associative learning capability in an Alzheimer disease (AD) C. elegans model. In summary, we report a novel function of spautin-1 in promoting mitophagy via the PINK1-PRKN pathway. As deficiency of mitophagy is closely implicated in the pathogenesis of neurodegenerative disorders, the pro-mitophagy function of spautin-1 might suggest its therapeutic potential in neurodegenerative disorders such as AD.Abbreviations: AD, Alzheimer disease; ATG, autophagy related; BafA1, bafilomycin A₁; CALCOCO2/NDP52, calcium binding and coiled-coil domain 2; CCCP, carbonyl cyanide m-chlorophenyl hydrazone; COX4/COX IV, cytochrome c oxidase subunit 4; EBSS, Earle's balanced salt; ECAR, extracellular acidification rate; GFP, green fluorescent protein; IA, isoamyl alcohol; IMM, inner mitochondrial membrane; MAP1LC3/LC3, microtubule associated protein 1 light chain 3; MMP, mitochondrial membrane potential; mtDNA, mitochondrial DNA; nDNA, nuclear DNA; O/A, oligomycin-antimycin; OCR, oxygen consumption rate; OMM, outer mitochondrial membrane; OPTN, optineurin; PARL, presenilin associated rhomboid like; PINK1, PTEN induced kinase 1; PRKN, parkin RBR E3 ubiquitin protein ligase; p-Ser65-Ub, phosphorylation of Ub at Ser65; TIMM23, translocase of inner mitochondrial membrane 23; TOMM, translocase of outer mitochondrial membrane; USP10, ubiquitin specific peptidase 10; USP13, ubiquitin specific peptidase 13; VAL, valinomycin; YFP, yellow fluorescent protein.

Macroautophagy/autophagy plays a crucial role in inhibiting viral replication and regulating the host's immune response. The autophagy receptor SQSTM1/p62 (sequestosome 1) restricts viral replication by directing specific viral proteins to phagophores for degradation. In this study, we investigate the reciprocal relationship between Zika virus (ZIKV) and selective autophagy mediated by SQSTM1/p62. We show that NS2B3 protease encoded by ZIKV cleaves human SQSTM1/p62 at arginine 265 (R265). This cleavage also occurs with endogenous SQSTM1 in ZIKV-infected cells. Furthermore, overexpression of SQSTM1 inhibits ZIKV replication in A549 cells, while its absence increases viral titer. We have also shown that SQSTM1 impedes ZIKV replication by interacting with NS3 and NS5 and directing them to autophagic degradation, and that NS2B3-mediated cleavage could potentially alter this antiviral function of SQSTM1. Taken together, our study highlights the role of SQSTM1-mediated selective autophagy in the host's antiviral defense against ZIKV and uncovers potential viral evasion strategies that exploit the host's autophagic machinery to ensure successful infection.Abbreviation: Cas9: CRISPR-associated protein 9; Co-IP: co-immunoprecipitation; CRISPR: clustered regularly interspaced short palindromic repeats; DENV: dengue virus; GFP: green fluorescent protein; IFA: indirect immunofluorescence assay; KIR: KEAP1-interacting region; KO: knockout; LIR: MAP1LC3/LC3-interacting region; mAb: monoclonal antibody; NBR1: NBR1 autophagy cargo receptor; OPTN: optineurin; pAb: polyclonal antibody; PB1: Phox/BEM1 domain; R265A, a SQSTM1 construct with the arginine (R) residue at position 265 replaced with glutamic acid (A); SQSTM1: sequestosome 1; SQSTM1-C, C-terminal fragment of SQSTM1; SQSTM1-N, N-terminal fragment of SQSTM1; SVV: Seneca Valley virus; TAX1BP1: Tax1 binding protein 1; TBD: TRAF6-binding domain; TCID₅₀: 50% tissue culture infective dose; UBA: ubiquitin-associated domain; Ub: ubiquitin; WT: wild type; ZIKV: Zika virus; ZZ: ZZ-type zinc finger domain.

Age-related macular degeneration (AMD) is the leading cause of blindness in the elderly, with the non-neovascular or atrophic form being the most common. Current treatment options are limited, emphasizing the urgent need for new therapeutic strategies. Our key finding is that increased levels of AKT2 in the RPE cells impair lysosomal function and trigger secretory autophagy; a non-canonical macroautophagy/autophagy pathway where cellular materials are released via the plasma membrane rather than being degraded by lysosomes. We showed that this process involves a protein complex, AKT2-SYTL1-TRIM16-SNAP23, releasing factors contributing to drusen biogenesis, a clinical hallmark of AMD development. Importantly, SIRT5 can inhibit this pathway, potentially offering a protective effect. Understanding mechanisms by which this non-canonical autophagy pathway promotes extracellular waste accumulation could provide new insights into drusen biogenesis. Future therapies for atrophic AMD could focus on regulating secretory autophagy or manipulating proteins involved in this process.

Macroautophagy/autophagy maintains cellular homeostasis by degrading cytoplasmic components and its disruption is linked to Parkinson disease (PD), which is characterized by dopamine depletion and the accumulation of SNCA/α-synuclein aggregates in neurons. Therefore, activation of autophagy is considered a therapeutic strategy for PD; however, autophagy inducers have not yet been developed as therapeutic drugs because they are involved in a wide range of signaling pathways. Here, we focused on the lysosomal clustering around the microtubule-organizing center (MTOC) that can regulate the process of autophagosome-lysosome fusion, the final step of autophagy, and examined how lysosomal clustering affects protein degradation through autophagy. Our study identified six compounds from a high-content screen of 1,200 clinically approved drugs that induce both lysosomal clustering and autophagy. Notably, albendazole reduced SNCA aggregates in a PD model by lysosomal clustering and autophagy. These findings suggest that targeting lysosomal clustering could offer new therapeutic insights for PD.

Reticulophagy, which directs the endoplasmic reticulum (ER) to the phagophore for sequestration within an autophagosome and subsequent lysosomal degradation via specific receptors, is essential for ER quality control and is implicated in various diseases. This study utilizes Drosophila to establish an in vivo model for reticulophagy. Starvation-induced reticulophagy is detected across multiple tissues in Drosophila. Whole-body upregulation or downregulation of the expression of reticulophagy receptors, atl and Rtnl1, negatively affects fly health. Notably, moderate upregulation of reticulophagy in neuronal tissues by overexpressing these receptors reduces age-related degeneration. In a Drosophila Alzheimer model expressing human APP (amyloid beta precursor protein), reticulophagy is compromised. Correcting reticulophagy by enhancing atl and Rtnl1 expression in the neurons promotes APP degradation, significantly reducing neurodegenerative symptoms. However, overexpression of mutated atl and Rtnl1, which disrupts the interaction of the corresponding proteins with Atg8, does not alleviate these symptoms, emphasizing the importance of receptor functionality. These findings support modulating reticulophagy as a therapeutic strategy for aging and neurodegenerative diseases associated with ER protein accumulation.