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Nature Jan 2024Digested dietary fats are taken up by enterocytes where they are assembled into pre-chylomicrons in the endoplasmic reticulum followed by transport to the Golgi for...
Digested dietary fats are taken up by enterocytes where they are assembled into pre-chylomicrons in the endoplasmic reticulum followed by transport to the Golgi for maturation and subsequent secretion to the circulation. The role of mitochondria in dietary lipid processing is unclear. Here we show that mitochondrial dysfunction in enterocytes inhibits chylomicron production and the transport of dietary lipids to peripheral organs. Mice with specific ablation of the mitochondrial aspartyl-tRNA synthetase DARS2 (ref. ), the respiratory chain subunit SDHA or the assembly factor COX10 (ref. ) in intestinal epithelial cells showed accumulation of large lipid droplets (LDs) in enterocytes of the proximal small intestine and failed to thrive. Feeding a fat-free diet suppressed the build-up of LDs in DARS2-deficient enterocytes, which shows that the accumulating lipids derive mostly from digested fat. Furthermore, metabolic tracing studies revealed an impaired transport of dietary lipids to peripheral organs in mice lacking DARS2 in intestinal epithelial cells. DARS2 deficiency caused a distinct lack of mature chylomicrons concomitant with a progressive dispersal of the Golgi apparatus in proximal enterocytes. This finding suggests that mitochondrial dysfunction results in impaired trafficking of chylomicrons from the endoplasmic reticulum to the Golgi, which in turn leads to storage of dietary lipids in large cytoplasmic LDs. Taken together, these results reveal a role for mitochondria in dietary lipid transport in enterocytes, which might be relevant for understanding the intestinal defects observed in patients with mitochondrial disorders.
Topics: Animals; Mice; Aspartate-tRNA Ligase; Chylomicrons; Dietary Fats; Electron Transport Complex II; Endoplasmic Reticulum; Enterocytes; Epithelial Cells; Golgi Apparatus; Intestines; Lipid Droplets; Lipid Metabolism; Mitochondria
PubMed: 38123683
DOI: 10.1038/s41586-023-06857-0 -
ELife Apr 2024Mapping proteins in and associated with the Golgi apparatus reveals how this cellular compartment emerges in budding yeast and progresses over time.
Mapping proteins in and associated with the Golgi apparatus reveals how this cellular compartment emerges in budding yeast and progresses over time.
Topics: Golgi Apparatus; Saccharomycetales
PubMed: 38629949
DOI: 10.7554/eLife.97430 -
Nature Methods Apr 2024Although StayGold is a bright and highly photostable fluorescent protein, its propensity for obligate dimer formation may hinder applications in molecular fusion and...
Although StayGold is a bright and highly photostable fluorescent protein, its propensity for obligate dimer formation may hinder applications in molecular fusion and membrane targeting. To attain monovalent as well as bright and photostable labeling, we engineered tandem dimers of StayGold to promote dispersibility. On the basis of the crystal structure of this fluorescent protein, we disrupted the dimerization to generate a monomeric variant that offers improved photostability and brightness compared to StayGold. We applied the new monovalent StayGold tools to live-cell imaging experiments using spinning-disk laser-scanning confocal microscopy or structured illumination microscopy. We achieved cell-wide, high-spatiotemporal resolution and sustained imaging of dynamic subcellular events, including the targeting of endogenous condensin I to mitotic chromosomes, the movement of the Golgi apparatus and its membranous derivatives along microtubule networks, the distribution of cortical filamentous actin and the remolding of cristae membranes within mobile mitochondria.
Topics: Mitochondria; Golgi Apparatus; Microtubules; Microscopy, Confocal
PubMed: 38036853
DOI: 10.1038/s41592-023-02085-6 -
Nature Nov 2023During nutrient stress, macroautophagy degrades cellular macromolecules, thereby providing biosynthetic building blocks while simultaneously remodelling the proteome....
During nutrient stress, macroautophagy degrades cellular macromolecules, thereby providing biosynthetic building blocks while simultaneously remodelling the proteome. Although the machinery responsible for initiation of macroautophagy has been well characterized, our understanding of the extent to which individual proteins, protein complexes and organelles are selected for autophagic degradation, and the underlying targeting mechanisms, is limited. Here we use orthogonal proteomic strategies to provide a spatial proteome census of autophagic cargo during nutrient stress in mammalian cells. We find that macroautophagy has selectivity for recycling membrane-bound organelles (principally Golgi and endoplasmic reticulum). Through autophagic cargo prioritization, we identify a complex of membrane-embedded proteins, YIPF3 and YIPF4, as receptors for Golgiphagy. During nutrient stress, YIPF3 and YIPF4 interact with ATG8 proteins through LIR motifs and are mobilized into autophagosomes that traffic to lysosomes in a process that requires the canonical autophagic machinery. Cells lacking YIPF3 or YIPF4 are selectively defective in elimination of a specific cohort of Golgi membrane proteins during nutrient stress. Moreover, YIPF3 and YIPF4 play an analogous role in Golgi remodelling during programmed conversion of stem cells to the neuronal lineage in vitro. Collectively, the findings of this study reveal prioritization of membrane protein cargo during nutrient-stress-dependent proteome remodelling and identify a Golgi remodelling pathway that requires membrane-embedded receptors.
Topics: Animals; Autophagy; Autophagy-Related Protein 8 Family; Endoplasmic Reticulum; Golgi Apparatus; Mammals; Membrane Proteins; Nutrients; Proteome; Proteomics
PubMed: 37757899
DOI: 10.1038/s41586-023-06657-6 -
Brain : a Journal of Neurology Nov 2023Moyamoya disease is an uncommon cerebrovascular disorder characterized by steno-occlusive changes in the circle of Willis and abnormal vascular network development. Ring...
Moyamoya disease is an uncommon cerebrovascular disorder characterized by steno-occlusive changes in the circle of Willis and abnormal vascular network development. Ring finger protein 213 (RNF213) has been identified as an important susceptibility gene for Asian patients, but researchers have not completely elucidated whether RNF213 mutations affect the pathogenesis of moyamoya disease. Using donor superficial temporal artery samples, whole-genome sequencing was performed to identify RNF213 mutation types in patients with moyamoya disease, and histopathology was performed to compare morphological differences between patients with moyamoya disease and intracranial aneurysm. The vascular phenotype of RNF213-deficient mice and zebrafish was explored in vivo, and RNF213 knockdown in human brain microvascular endothelial cells was employed to analyse cell proliferation, migration and tube formation abilities in vitro. After bioinformatics analysis of both cell and bulk RNA-seq data, potential signalling pathways were measured in RNF213-knockdown or RNF213-knockout endothelial cells. We found that patients with moyamoya disease carried pathogenic mutations of RNF213 that were positively associated with moyamoya disease histopathology. RNF213 deletion exacerbated pathological angiogenesis in the cortex and retina. Reduced RNF213 expression led to increased endothelial cell proliferation, migration and tube formation. Endothelial knockdown of RNF213 activated the Hippo pathway effector Yes-associated protein (YAP)/tafazzin (TAZ) and promoted the overexpression of the downstream effector VEGFR2. Additionally, inhibition of YAP/TAZ resulted in altered cellular VEGFR2 distribution due to defects in trafficking from the Golgi apparatus to the plasma membrane and reversed RNF213 knockdown-induced angiogenesis. All these key molecules were validated in ECs isolated from RNF213-deficient animals. Our findings may suggest that loss-of-function of RNF213 mediates the pathogenesis of moyamoya disease via the Hippo pathway.
Topics: Humans; Animals; Mice; Moyamoya Disease; Endothelial Cells; Hippo Signaling Pathway; Zebrafish; Neovascularization, Pathologic; Genetic Predisposition to Disease; Adenosine Triphosphatases; Ubiquitin-Protein Ligases
PubMed: 37399508
DOI: 10.1093/brain/awad225 -
Cell Reports Jun 2023The nuclear factor κB (NF-κB) pathway plays essential roles in innate and adaptive immunity, but little is known how NF-κB signaling is compartmentalized and...
The nuclear factor κB (NF-κB) pathway plays essential roles in innate and adaptive immunity, but little is known how NF-κB signaling is compartmentalized and spatiotemporally activated in the cytoplasm. Here, we show that the lipogenesis signal cascade Scap-SREBP1-S1P/S2P orchestrates the homeostasis and spatiotemporal activation of NF-κB. SREBP cleavage-activating protein (Scap) and sterol regulatory element-binding protein 1 (SREBP1) form a super complex with inhibitors of NF-κB α (IκBα) to associate NF-κB close to the endoplasmic reticulum (ER). Upon lipopolysaccharide (LPS) stimulation, Scap transports the complex to the Golgi apparatus, where SREBP1 is cleaved by site-1 protease (S1P)/S2P, liberating IκBα for IκB kinase (Ikk)-mediated phosphorylation and subsequent activation of NF-κB. Loss of Scap or inhibition of S1P or S2P diminishes, while SREBP1 deficiency augments, LPS-induced NF-κB activation and subsequent inflammatory responses. Our results reveal the Scap-SREBP1 complex as an additional cytoplasmic checkpoint for NF-κB homeostasis and unveil the Golgi apparatus as the optimal cellular platform for NF-κB activation, providing insights into the crosstalk between lipogenesis signaling and immunity.
Topics: Homeostasis; Lipogenesis; Lipopolysaccharides; NF-kappa B; NF-KappaB Inhibitor alpha; Sterol Regulatory Element Binding Protein 1; Humans; Animals; Mice
PubMed: 37267109
DOI: 10.1016/j.celrep.2023.112586 -
Nature Cell Biology Dec 2023Drugs that selectively kill senescent cells (senolytics) improve the outcomes of cancer, fibrosis and age-related diseases. Despite their potential, our knowledge of the...
Drugs that selectively kill senescent cells (senolytics) improve the outcomes of cancer, fibrosis and age-related diseases. Despite their potential, our knowledge of the molecular pathways that affect the survival of senescent cells is limited. To discover senolytic targets, we performed RNAi screens and identified coatomer complex I (COPI) vesicle formation as a liability of senescent cells. Genetic or pharmacological inhibition of COPI results in Golgi dispersal, dysfunctional autophagy, and unfolded protein response-dependent apoptosis of senescent cells, and knockdown of COPI subunits improves the outcomes of cancer and fibrosis in mouse models. Drugs targeting COPI have poor pharmacological properties, but we find that N-myristoyltransferase inhibitors (NMTi) phenocopy COPI inhibition and are potent senolytics. NMTi selectively eliminated senescent cells and improved outcomes in models of cancer and non-alcoholic steatohepatitis. Our results suggest that senescent cells rely on a hyperactive secretory apparatus and that inhibiting trafficking kills senescent cells with the potential to treat various senescence-associated diseases.
Topics: Mice; Animals; Senotherapeutics; Golgi Apparatus; Cellular Senescence; Neoplasms; Fibrosis
PubMed: 38012402
DOI: 10.1038/s41556-023-01287-6 -
The Journal of Clinical Investigation Aug 2023CXCR7 is an atypical chemokine receptor that recruits β-arrestin (ARRB2) and internalizes into clathrin-coated intracellular vesicles where the complex acts as a...
CXCR7 is an atypical chemokine receptor that recruits β-arrestin (ARRB2) and internalizes into clathrin-coated intracellular vesicles where the complex acts as a scaffold for cytoplasmic kinase assembly and signal transduction. Here, we report that CXCR7 was elevated in the majority of prostate cancer (PCa) cases with neuroendocrine features (NEPC). CXCR7 markedly induced mitotic spindle and cell cycle gene expression. Mechanistically, we identified Aurora Kinase A (AURKA), a key regulator of mitosis, as a novel target that was bound and activated by the CXCR7-ARRB2 complex. CXCR7 interacted with proteins associated with microtubules and golgi, and, as such, the CXCR7-ARRB2-containing vesicles trafficked along the microtubules to the pericentrosomal golgi apparatus, where the complex interacted with AURKA. Accordingly, CXCR7 promoted PCa cell proliferation and tumor growth, which was mitigated by AURKA inhibition. In summary, our study reveals a critical role of CXCR7-ARRB2 in interacting and activating AURKA, which can be targeted by AURKA inhibitors to benefit a subset of patients with NEPC.
Topics: Male; Humans; Aurora Kinase A; Signal Transduction; Receptors, CXCR; Prostatic Neoplasms; Cell Proliferation; Cell Line, Tumor
PubMed: 37347559
DOI: 10.1172/JCI166248