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Autophagy Jun 2024In neurons, macroautophagy/autophagy is a frequent and critical process. In the axon, autophagy begins in the axon terminal, where most nascent autophagosomes form....
In neurons, macroautophagy/autophagy is a frequent and critical process. In the axon, autophagy begins in the axon terminal, where most nascent autophagosomes form. After formation, autophagosomes must initiate transport to exit the axon terminal and move toward the cell body via retrograde transport. During retrograde transport these autophagosomes mature through repetitive fusion events. Complete lysosomal cargo degradation occurs largely in the cell body. The precipitating events to stimulate retrograde autophagosome transport have been debated but their importance is clear: disrupting neuronal autophagy or autophagosome transport is detrimental to neuronal health and function. We have identified the HOPS complex as essential for early autophagosome maturation and consequent initiation of retrograde transport from the axon terminal. In yeast and mammalian cells, HOPS controls fusion between autophagosomes and late endosomes with lysosomes. Using zebrafish strains with loss-of-function mutations in and , core components of the HOPS complex, we found that disruption of HOPS eliminates autophagosome maturation and disrupts retrograde autophagosome transport initiation from the axon terminal. We confirmed this phenotype was due to loss of HOPS complex formation using an endogenous deletion of the HOPS binding domain in Vps18. Finally, using pharmacological inhibition of lysosomal proteases, we show that initiation of autophagosome retrograde transport requires autophagosome maturation. Together, our data demonstrate that HOPS-mediated fusion events are critical for retrograde autophagosome transport initiation through promoting autophagosome maturation. This reveals critical roles for the HOPS complex in neuronal autophagy which deepens our understanding of the cellular pathology of HOPS-complex linked neurodegenerative diseases.: CORVET: Class C core vacuole/endosome tethering; gRNA: guide RNA; HOPS: homotypic fusion and protein sorting; pLL: posterior lateral line; Vps18: VPS18 core subunit of CORVET and HOPS complexes; Vps41: VPS41 subunit of HOPS complex.
PubMed: 38899385
DOI: 10.1080/15548627.2024.2366122 -
Nature Communications Jun 2024Autophagy is relevant for diverse processes in eukaryotic cells, making its regulation of fundamental importance. The formation and maturation of autophagosomes require...
Autophagy is relevant for diverse processes in eukaryotic cells, making its regulation of fundamental importance. The formation and maturation of autophagosomes require a complex choreography of numerous factors. The endosomal sorting complex required for transport (ESCRT) is implicated in the final step of autophagosomal maturation by sealing of the phagophore membrane. ESCRT-III components were shown to mediate membrane scission by forming filaments that interact with cellular membranes. However, the molecular mechanisms underlying the recruitment of ESCRTs to non-endosomal membranes remain largely unknown. Here we focus on the ESCRT-associated protein ALG2-interacting protein X (ALIX) and identify Ca-dependent lipid binding protein 1 (CaLB1) as its interactor. Our findings demonstrate that CaLB1 interacts with AUTOPHAGY8 (ATG8) and PI(3)P, a phospholipid found in autophagosomal membranes. Moreover, CaLB1 and ALIX localize with ATG8 on autophagosomes upon salt treatment and assemble together into condensates. The depletion of CaLB1 impacts the maturation of salt-induced autophagosomes and leads to reduced delivery of autophagosomes to the vacuole. Here, we propose a crucial role of CaLB1 in augmenting phase separation of ALIX, facilitating the recruitment of ESCRT-III to the site of phagophore closure thereby ensuring efficient maturation of autophagosomes.
Topics: Arabidopsis; Autophagosomes; Endosomal Sorting Complexes Required for Transport; Arabidopsis Proteins; Calcium-Binding Proteins; Autophagy; Phosphatidylinositol Phosphates; Autophagy-Related Protein 8 Family; Vacuoles; Phase Separation
PubMed: 38898014
DOI: 10.1038/s41467-024-49485-6 -
Journal of Hazardous Materials Jun 2024Micro- and nanoplastic pollution has emerged as a significant global concern due to their extensive presence in the environment and potential adverse effects on human...
Micro- and nanoplastic pollution has emerged as a significant global concern due to their extensive presence in the environment and potential adverse effects on human health. Nanoplastics can enter the human circulatory system and accumulate in the liver, disrupting hepatic metabolism and causing hepatotoxicity. However, the precise mechanism remains uncertain. Lipophagy is an alternative mechanism of lipid metabolism involving autophagy. This study aims to explore how polystyrene nanoplastics (PSNPs) influence lipid metabolism in hepatocytes via lipophagy. Initially, it was found that PSNPs were internalized by human hepatocytes, resulting in decreased cell viability. PSNPs were found to induce the accumulation of lipid droplets (LDs), with autophagy inhibition exacerbating this accumulation. Then, PSNPs were proved to activate lipophagy by recruiting LDs into autophagosomes and block the lipophagic flux by impairing lysosomal function, inhibiting LD degradation. Ultimately, PSNPs were shown to activate lipophagy through the AMPK/ULK1 pathway, and knocking down AMPK exacerbated lipid accumulation in hepatocytes. Overall, these results indicated that PSNPs triggered lipophagy via the AMPK/ULK1 pathway and blocked lipophagic flux, leading to lipid accumulation in hepatocytes. Thus, this study identifies a novel mechanism underlying nanoplastic-induced lipid accumulation, providing a foundation for the toxicity study and risk assessments of nanoplastics.
PubMed: 38897115
DOI: 10.1016/j.jhazmat.2024.134878 -
International Journal of Molecular... Jun 2024Sprouty-related enabled/vasodilator-stimulated phosphoprotein homology 1 domain containing 2 (SPRED2) is an inhibitor of the mitogen-activated protein kinase...
Sprouty-related enabled/vasodilator-stimulated phosphoprotein homology 1 domain containing 2 (SPRED2) is an inhibitor of the mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) pathway and has been shown to promote autophagy in several cancers. Here, we aimed to determine whether SPRED2 plays a role in autophagy in hepatocellular carcinoma (HCC) cells. The Cancer Genome Atlas (TCGA) Liver Cancer Database showed a negative association between the level of SPRED2 and p62, a ubiquitin-binding scaffold protein that accumulates when autophagy is inhibited. Immunohistochemically, accumulation of p62 was detected in human HCC tissues with low SPRED2 expression. Overexpression of SPRED2 in HCC cells increased the number of autophagosomes and autophagic vacuoles containing damaged mitochondria, decreased p62 levels, and increased levels of light-chain-3 (LC3)-II, an autophagy marker. In contrast, SPRED2 deficiency increased p62 levels and decreased LC3-II levels. SPRED2 expression levels were negatively correlated with translocase of outer mitochondrial membrane 20 (TOM20) expression levels, suggesting its role in mitophagy. Mechanistically, SPRED2 overexpression reduced ERK activation followed by the mechanistic or mammalian target of rapamycin complex 1 (mTORC1)-mediated signaling pathway, and SPRED2 deficiency showed the opposite pattern. Finally, hepatic autophagy was impaired in the liver of SPRED2-deficient mice with hepatic lipid droplet accumulation in response to starvation. These results indicate that SPRED2 is a critical regulator of autophagy not only in HCC cells, but also in hepatocytes, and thus the manipulation of this process may provide new insights into liver pathology.
Topics: Humans; Carcinoma, Hepatocellular; Liver Neoplasms; Autophagy; Hepatocytes; Animals; Mice; Cell Line, Tumor; Mechanistic Target of Rapamycin Complex 1; MAP Kinase Signaling System; Mitophagy; Repressor Proteins
PubMed: 38892460
DOI: 10.3390/ijms25116269 -
International Journal of Molecular... Jun 2024This study demonstrated the anticancer efficacy of chalcones with indole moiety (MIPP, MOMIPP) in fibrosarcoma cells for the first time. The results showed that MIPP and...
This study demonstrated the anticancer efficacy of chalcones with indole moiety (MIPP, MOMIPP) in fibrosarcoma cells for the first time. The results showed that MIPP and MOMIPP reduced the viability of HT-1080 cells in a concentration-dependent manner. MOMIPP was more active than MIPP in HT-1080 cells, showing lower IC values (3.67 vs. 29.90 μM). Both compounds at a concentration of 1 μM induced apoptosis in HT-1080 cells, causing death strictly related to caspase activation, as cell viability was restored when the caspase inhibitor Z-VAD was added. Reactive oxygen species production was approximately 3-fold higher than in control cells, and cotreatment with the inhibitor of mitochondrial ATPase oligomycin diminished this effect. Such effects were also reflected in mitochondrial dysfunction, including decreased membrane potential. Interestingly, the compounds that were studied caused massive vacuolization in HT-1080 cells. Immunocytochemical staining and TEM analysis showed that HT-1080 cells exhibited increased expression of the LC3-II protein and the presence of autophagosomes with a double membrane, respectively. Both compounds induced apoptosis, highlighting a promising link between autophagy and apoptosis. This connection could be a new target for therapeutic strategies to overcome chemoresistance, which is a significant cause of treatment failure and tumour recurrence in fibrosarcoma following traditional chemotherapy.
Topics: Humans; Apoptosis; Fibrosarcoma; Autophagy; Indoles; Cell Line, Tumor; Reactive Oxygen Species; Chalcones; Membrane Potential, Mitochondrial; Cell Survival; Antineoplastic Agents; Mitochondria
PubMed: 38892288
DOI: 10.3390/ijms25116100 -
International Journal of Molecular... May 2024Periodontitis development arises from the intricate interplay between bacterial biofilms and the host's immune response, where macrophages serve pivotal roles in defense...
Periodontitis development arises from the intricate interplay between bacterial biofilms and the host's immune response, where macrophages serve pivotal roles in defense and tissue homeostasis. Here, we uncover the mitigative effect of copper chelator Tetrathiomolybdate (TTM) on periodontitis through inhibiting cuproptosis, a newly identified form of cell death which is dependent on copper. Our study reveals concurrent cuproptosis and a macrophage marker within murine models. In response to lipopolysaccharide (LPS) stimulation, macrophages exhibit elevated cuproptosis-associated markers, which are mitigated by the administration of TTM. TTM treatment enhances autophagosome expression and mitophagy-related gene expression, countering the LPS-induced inhibition of autophagy flux. TTM also attenuates the LPS-induced fusion of autophagosomes and lysosomes, the degradation of lysosomal acidic environments, lysosomal membrane permeability increase, and cathepsin B secretion. In mice with periodontitis, TTM reduces cuproptosis, enhances autophagy flux, and decreases levels. Our findings underscore the crucial role of copper-chelating agent TTM in regulating the cuproptosis/mitophagy/lysosome pathway during periodontitis inflammation, suggesting TTM as a promising approach to alleviate macrophage dysfunction. Modulating cuproptosis through TTM treatment holds potential for periodontitis intervention.
Topics: Animals; Lysosomes; Mice; Periodontitis; Autophagy; Molybdenum; Copper; Chelating Agents; Lipopolysaccharides; Macrophages; Chelation Therapy; Inflammation; Mice, Inbred C57BL; Male
PubMed: 38892077
DOI: 10.3390/ijms25115890 -
Frontiers in Cell and Developmental... 2024Leukemia is a life-threatening malignant tumor of the hematopoietic system. Currently, the main treatment modalities are chemotherapy and hematopoietic stem cell... (Review)
Review
Leukemia is a life-threatening malignant tumor of the hematopoietic system. Currently, the main treatment modalities are chemotherapy and hematopoietic stem cell transplantation. However, increased drug resistance due to decreased sensitivity of leukemia cells to chemotherapeutic drugs presents a major challenge in current treatments. Autophagy-associated proteins involved in autophagy initiation have now been shown to be involved in the development of various types of leukemia cells and are associated with drug resistance. Therefore, this review will explore the roles of autophagy-related proteins involved in four key autophagic processes: induction of autophagy and phagophore formation, phagophore extension, and autophagosome formation, on the development of various types of leukemias as well as drug resistance. Autophagy may become a promising therapeutic target for treating leukemia.
PubMed: 38887520
DOI: 10.3389/fcell.2024.1394140 -
Current Medicinal Chemistry Jun 2024Oral squamous cell carcinoma (OSCC) represents the primary form of oral cancer, posing a significant global health threat. The existing chemotherapy options are...
BACKGROUND
Oral squamous cell carcinoma (OSCC) represents the primary form of oral cancer, posing a significant global health threat. The existing chemotherapy options are accompanied by notable side effects impacting patient treatment adherence. Consequently, the exploration and development of novel substances with enhanced anticancer effects and fewer side effects have become pivotal in the realms of biological and chemical science.
OBJECTIVE
This work presents the pioneering examples of naphthoquinone-coumarin hybrids as a new category of highly effective cytotoxic substances targeting oral squamous cell carcinoma (OSCC).
METHODS
Given the significance of both naphthoquinones and coumarins as essential pharmacophores/ privileged structures in the quest for anticancer compounds, this study focused on the synthesis and evaluation of novel naphthoquinones/coumarin hybrids against oral squamous cell carcinoma.
RESULTS
By several in vitro, in silico, and in vivo approaches, we demonstrated that compound 6e was highly cytotoxic against OSCC cells and several other cancer cell types and was more selective than current chemotherapeutic drugs (carboplatin) and the naphthoquinone lapachol. Furthermore, compound 6e was non-hemolytic and tolerated in vivo at 50 mg/kg with an LD50 of 62.5 mg/kg. Furthermore, compound 6e did not induce apoptosis and cell cycle arrest but led to intracellular vesicle formation with LC3 aggregation in autophagosomes, suggesting an autophagic cell death. Additionally, 6e had a high-affinity potential for PKM2 protein, higher than the known ligands, such as lapachol or shikonin, and was able to inhibit this enzyme activity in vitro.
CONCLUSION
We assert that compound 6e shows promise as a potential lead for a novel chemotherapeutic drug targeting OSCC, with potential applicability to other cancer types.
PubMed: 38877863
DOI: 10.2174/0109298673298471240605072658 -
Alzheimer's Research & Therapy Jun 2024Our study aims to evaluate the genetic and phenotypic spectrum of Frontotemporal dementia (FTD) gene variant carriers in Chinese populations, investigate mutation...
BACKGROUND
Our study aims to evaluate the genetic and phenotypic spectrum of Frontotemporal dementia (FTD) gene variant carriers in Chinese populations, investigate mutation frequencies, and assess the functional properties of TBK1 and OPTN variants.
METHODS
Clinically diagnosed FTD patients underwent genetic analysis through exome sequencing, repeat-primed polymerase chain reaction, and Sanger sequencing. TBK1 and OPTN variants were biologically characterized in vitro using immunofluorescence, immunoprecipitation, and immunoblotting analysis. The frequencies of genes implicated in FTD in China were analyzed through a literature review and meta-analysis.
RESULTS
Of the 261 Chinese FTD patients, 61 (23.4%) carried potential causative variants in FTD-related genes, including MAPT (n = 17), TBK1 (n = 7), OPTN (n = 6), GRN (n = 6), ANXA11 (n = 4), CHMP2B (n = 3), C9orf72 GGGGCC repeats (n = 2), CYLD (n = 2), PRNP (n = 2), SQSTM1 (n = 2), TARDBP (n = 2), VCP (n = 1), CCNF (n = 1), CHCHD10 (n = 1), SIGMAR1 (n = 1), CHCHD2 (n = 1), FUS (n = 1), TMEM106B (n = 1), and UBQLN2 (n = 1). 29 variants can be considered novel, including the MAPT p.D54N, p.E342K, p.R221P, p.T263I, TBK1 p.E696G, p.I37T, p.E232Q, p.S398F, p.T78A, p.Q150P, p.W259fs, OPTN p.R144G, p.F475V, GRN p.V473fs, p.C307fs, p.R101fs, CHMP2B p.K6N, p.R186Q, ANXA11 p.Q155*, CYLD p.T157I, SQSTM1 p.S403A, UBQLN2 p.P509H, CCNF p.S160N, CHCHD10 p.A8T, SIGMAR1 p.S117L, CHCHD2 p.P53fs, FUS p.S235G & p.S236G, and TMEM106B p.L144V variants. Patients with TBK1 and OPTN variants presented with heterogeneous clinical phenotypes. Functional analysis demonstrated that TBK1 I37T and E232Q mutants showed decreased autophosphorylation, and the OPTN phosphorylation was reduced by the TBK1 I37T mutant. The OPTN-TBK1 complex formation was enhanced by the TBK1 E696G mutant, while OPTN R144G and F475V mutants exhibited reduced recruitment to autophagosomes compared to the wild-type. The overall frequency of TBK1 and OPTN in Chinese FTD patients was 2.0% and 0.3%, respectively.
CONCLUSIONS
Our study demonstrates the extensive genetic and phenotypic heterogeneity of Chinese FTD patients. TBK1 mutations are the second most frequent cause of clinical FTD after MAPT in the Chinese.
Topics: Aged; Female; Humans; Male; Middle Aged; Cell Cycle Proteins; China; East Asian People; Frontotemporal Dementia; Genetic Predisposition to Disease; Membrane Transport Proteins; Mutation; Protein Serine-Threonine Kinases; Transcription Factor TFIIIA
PubMed: 38872230
DOI: 10.1186/s13195-024-01493-w -
The Journal of Clinical Investigation Jun 2024The identification of genes that confer either extension of lifespan or accelerate age-related decline was a step forward in understanding the mechanisms of ageing and...
The identification of genes that confer either extension of lifespan or accelerate age-related decline was a step forward in understanding the mechanisms of ageing and revealed that it is partially controlled by genetics and transcriptional programs. Here we discovered that the human DNA sequence C16ORF70 encoded for a protein, named MYTHO (Macroautophagy and YouTH Optimizer), which controls life- and health-span. MYTHO protein is conserved from C. elegans to humans and its mRNA was upregulated in aged mice and elderly people. Deletion of the ortholog myt-1 gene in C. elegans dramatically shortened lifespan and decreased animal survival upon exposure to oxidative stress. Mechanistically, MYTHO is required for autophagy likely because it acts as a scaffold that binds WIPI2 and BCAS3 to recruit and assemble the conjugation system at the phagophore, the nascent autophagosome. We conclude that MYTHO is a transcriptionally regulated initiator of autophagy that is central in promoting stress resistance and healthy ageing.
PubMed: 38869949
DOI: 10.1172/JCI165814