Autophagy最新文献

Mitochondrial damage in fibroblast-like synoviocytes (FLSs) is a key factor involved in the development and progression of rheumatoid arthritis (RA). In this study, we investigated the role of mitochondrial dysfunction of FLSs in the pathogenesis of RA. We induced inflammation by stimulating FLSs with TNF and IL17. Then, we transplanted fresh mitochondria into stimulated FLSs and evaluated the mitochondrial and lysosomal functions, macroautophagic/autophagic activity, and the STING1-associated cell death pathway. Next, we transplanted mitochondria or gold nanoparticle-conjugated mitochondria (GNP-Mito) into collagen-induced arthritis (CIA) mice and evaluated their therapeutic effects in vivo. Mitochondrial and lysosomal activities were decreased and autophagosomes accumulated in the stimulated FLSs. Furthermore, the STING1 signaling pathway and STING1-associated cell death were increased in the inflammatory condition. Mitochondrial transplantation into stimulated FLSs enhanced the mitochondrial and lysosomal activities and activated the autophagic activity, as demonstrated by decreased numbers of autophagosomes and increased numbers of autolysosomes. Mitochondrial transplantation decreased and increased the T_h17 and T_reg populations, respectively. Mitochondrial function and autophagic activity were enhanced by mitochondrial transplantation. Taken together, our results demonstrate that mitochondrial dysfunction in FLSs plays a pivotal role in the pathophysiology of RA and mitochondrial transplantation has therapeutic potential for RA development and progression.Abbreviations: ATP:adenosine triphosphate; CGAS: cyclic GMP-AMP synthase; CIA:collagen-induced arthritis; FLS: fibroblast-like synoviocytes; GNP:gold nanoparticle; ROS: reactive oxygen species; SQSTM1/p62:sequestosome 1; STING1: stimulator of interferon response cGAMPinteractor 1; MAP1LC3B/LC3B: microtubule associated protein 1 lightchain 3 beta.

Golgi fragmentation is a prominent early hallmark of neurodegenerative diseases such as Alzheimer disease (AD) and amyotrophic lateral sclerosis (ALS), yet the shared molecular mechanisms underlying this phenomenon remain poorly understood. Here we identify the E3 ubiquitin ligase ITCH as a central regulator of Golgi integrity and proteostasis. Elevated ITCH disrupts both cis- and trans-Golgi networks, dislocates lysosomal hydrolase sorting factors, and impairs maturation of hydrolases. The ensuing lysosomal dysfunction leads to autophagosome accumulation and defective clearance of accumulated cytoplasmic toxic proteins like TARDBP/TDP-43. Genetic and pharmacological inhibition of ITCH restores autolysosomal degradation and protects neurons in both mammalian and Drosophila models. Aberrant buildup of the deubiquitinase USP11 drives ITCH accumulation, intensifying neuronal proteotoxic stress in individuals with AD and ALS. These findings reveal a mechanistic pathway connecting Golgi disorganization, autolysosomal impairment, and proteotoxic stress in neurodegeneration.

TFEB (transcription factor EB) is a critical regulator of lysosomal biogenesis, macroautophagy/autophagy and energy homeostasis through controlling expression of genes belonging to the coordinated lysosomal expression and regulation network. AMP-activated protein kinase (AMPK) has been reported to phosphorylate TFEB at three conserved C-terminal serine residues (S466, S467, S469) and these phosphorylation events were reported to be essential for transcriptional activation of TFEB. In sharp contrast to this proposition, we demonstrate that AMPK activation leads to the dephosphorylation of the C-terminal sites. We show that a synthetic peptide encompassing the C-terminal serine residues of TFEB is a poor substrate of AMPK in vitro. Treatment of cells with an AMPK activator (MK-8722), glucose deprivation or MTOR inhibitor (torin1) robustly dephosphorylated TFEB not only at the MTORC1-targeted N-terminal serine sites, but also at the C-terminal sites. Loss of function of AMPK abrogated MK-8722- but not torin1-induced dephosphorylation and induction of the TFEB target genes.

NEU (neuraminidase) is a potential cross-reactive antigen for developing broadly protective influenza vaccine, but has suboptimal immunogenicity. We here report that, when NEU antigen was redirected into phagophores, and subsequently autophagosomes, by fusing with MAP1LC3B/LC3B (microtubule associated protein 1 light chain 3 beta; NEU-LC3B), it could efficiently activate the autophagosome-lysosome-major histocompatibility complex class II (MHC II) compartment pathway, and thus substantially improve the magnitude, breadth, and polyfunctionality of NEU-specific T cell immunity in mice. Remarkably, we identified several novel NEU-specific T-cell epitopes in response to NEU-LC3B-based immunization. Furthermore, mice immunized with NEU-based constructs were challenged with homologous A/CA/04/09 (H1N1), heterologous within-subtype strain A/Puerto Rico/8/1934 (PR8) (H1N1), and heterosubtypic A/Aichi/2/1968 (H3N2) virus, and the results demonstrated that NEU-LC3B-based vaccine provided a sterilizing immunity to homologous strains and cross-protection against antigenically distinct heterosubtypic challenge. In addition, cell depletion experiment demonstrated that T-cell-mediated immunity contributed to the NEU-LC3B-mediated immune protection. Collectively, this engineered NEU antigen with optimal immunogenicity represents a promising strategy for developing broadly protective influenza vaccines.

Previous studies have shown that SIGMAR1/Sigma-1 receptor (sigma non-opioid intracellular receptor 1) provides protective effects against lipopolysaccharide (LPS)-induced acute lung injury (ALI), however the underlying mechanism remains unclear. A recent study highlighted SIGMAR1's protective role against ferroptosis but did not fully elucidate the mechanism involved. Endothelial ferroptosis, which significantly affects microvascular permeability, has garnered increasing attention in research. In this context, we aimed to investigate how SIGMAR1 mitigates endothelial ferroptosis in ALI induced by LPS. PRE-084 (SIGMAR1 activator) inhibited endothelial ferroptosis and microvascular hyperpermeability in ALI induced by LPS; however, this effect was blocked by mitophagy inhibition. Knockout of sigmar1 worsened microvascular hyperpermeability and endothelial ferroptosis, but these effects were mitigated by activating SIRT3 (sirtuin 3). Conversely, inhibiting SIRT3 blocked the upregulation of SIGMAR1-mediated mitophagy and limited endothelial ferroptosis in ALI induced by LPS. In addition, LPS exposure led to the acetylation of lysine 498 in ATP5F1A/ATP5A1 (ATP synthase F1 subunit alpha). Importantly, downregulating ATP5F1A acetylation prevented the SIRT3 inhibition from blocking the effects of SIGMAR1 in facilitating mitophagy and preventing ferroptosis. Interestingly, downregulating ATP5F1A acetylation or activation of SIRT3 did not alter the effects of PRE-084 on ALI when mitophagy was inhibited, suggesting that SIGMAR1's ALI protective effects involve ATP5F1A- or SIRT3-dependent mitophagy. In conclusion, our findings indicate that SIGMAR1 alleviates endothelial ferroptosis and microvascular hyperpermeability in LPS-induced ALI through SIRT3-mediated mitophagy. Furthermore, the deacetylation of ATP5F1A at lysine 498 by SIRT3 is essential for SIGMAR1-mediated PRKN/parkindependent mitophagy.

The targeted degradation of oncogenic or misfolded proteins has emerged as a promising therapeutic strategy. While proteolysis-targeting chimeras (PROTACs) and related technologies have successfully hijacked the ubiquitin-proteasome system to eliminate disease-driving proteins, recent advances highlight the lysosome as a powerful alternative degradation route. Lysosome-based degradation strategies offer broader substrate scope, subcellular targeting flexibility, and the ability to degrade proteins beyond the reach of the proteasome. In this review, we provide a comprehensive overview of synthetic molecules and engineered systems designed to traffic target proteins to the lysosome. These include lysosome targeting chimeras (LYTACs), autophagy-targeting chimeras (AUTACs), autophagy-tethering compounds (ATTECs), and other modalities that exploit endogenous trafficking pathways for selective protein clearance. By mapping the current landscape of lysosome-targeting degraders, this article underscores the therapeutic potential of lysosomal proteolysis and outlines future directions for molecular engineering in this rapidly evolving field.

Despite the clinical success of PDCD1/PD-1 and CD274/PD-L1 immune checkpoint blockade in multiple cancers, its efficacy in colorectal cancer (CRC) remains limited. Here, we report that the combination of the tyrosine kinase inhibitor regorafenib with PDCD1 blockade enhances anti-tumor immunity in CRC, both in clinical observations and preclinical models. Mechanistically, regorafenib acts as a molecular glue, directly promoting the interaction between CD274 and the selective autophagy receptor SQSTM1/p62, leading to SQSTM1-mediated autophagic degradation of CD274 and restoration of T cell-mediated cytotoxicity. In summary, these findings identify a previously unrecognized role of regorafenib in modulating tumor immune evasion and provide a mechanistic rationale for its combination with PDCD1 inhibitors in CRC treatment.

PINK1-dependent activation of PRKN/parkin on depolarized mitochondria causes mitophagy. The deficiency of NME3, a nucleoside diphosphate kinase/NDPK on the outer mitochondria membrane (OMM), is associated with a fatal neurodegenerative disorder. Here, we report that NME3 deficiency impairs p-S65-ubiquitin (Ub)-dependent PRKN binding on depolarized mitochondria without involving the loss of Ub phosphorylation by PINK1. Our mechanistic investigation revealed that NME3 interacts with PLD6/MitoPLD to generate phosphatidic acid (PA) from cardiolipin on the OMM of damaged mitochondria after depolarization. This lipid signal is essential for positioning MFN2 nearby PINK1 for phosphorylation of Ub conjugates on MFN2, thus enabling the subsequent amplification of PRKN binding to mitochondria. We provide further evidence that mitochondria-endoplasmic reticulum (Mito-ER) tethering prohibits the proximity of MFN2 with PINK1 and PRKN amplification on mitochondria. Importantly, the loss of NME3-regulated PA signal causes Mito-ER tethering. Overall, our findings suggest that NME3 cooperates with PLD6 to generate PA as a critical step in Mito-ER untethering, allowing MFN2 access to PINK1 for p-S65-poly-Ub-dependent feedforward activation of PRKN.Abbreviation ACTB: actin beta; BDNF brain derived neurotrophic factor; CL: cardiolipin; CRISPR: clustered regularly interspaced short palindromic repeats; DAG: diacylglycerol; ER: endoplasmic reticulum; FCCP: carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone; FRET: Förster resonance energy transfer; IF: immunofluorescence; KO: knockout; KD: knockdown; LPIN1: lipin 1; MERCS: mitochondria-endoplasmic reticulum contact sites; MFN2: mitofusin 2; Mito: mitochondria; OMM: outer mitochondrial membrane; p-Ub: phosphorylated ubiquitin; PA: phosphatidic acid; PD: Parkinson disease; PINK1: PTEN induced kinase 1; PLA: proximity ligation assay; PLD6/MitoPLD: phospholipase D family member 6; PRKN: parkin RBR E3 ubiquitin protein ligase; RA: retinoic acid; RT-qPCR: reverse transcription-quantitative polymerase chain reaction; TEM: transmission electron microscopy; TN-NME3: TOMM20-NΔ-NME3; TOMM20: translocase of outer mitochondrial membrane 20; TUBB: tubulin beta class I; Ub: ubiquitin; VDAC: voltage dependent anion channel; WB: western blot.