EMBO Reports最新文献

Pro-inflammatory macrophage activation is a hallmark example of how mitochondria serve as signaling organelles. Oxidative phosphorylation sharply decreases upon classical macrophage activation, as mitochondria are thought to shift from ATP production towards accumulating signals that amplify effector function. However, evidence is conflicting regarding whether this collapse in respiration is essential or dispensable. Here we systematically examine this question and show that reduced oxidative phosphorylation is not required for pro-inflammatory macrophage activation. Different pro-inflammatory stimuli elicit varying effects on bioenergetic parameters, and pharmacologic and genetic models of electron transport chain inhibition show no causative link between respiration and macrophage activation. Furthermore, the signaling metabolites succinate and itaconate can accumulate independently of characteristic breaks in the TCA cycle in mouse and human macrophages, and peritoneal macrophages can be activated in vivo without inhibition of oxidative phosphorylation. The results indicate there is plasticity in the metabolic phenotypes that can support pro-inflammatory macrophage activation.

Recent genetic studies by the Schizophrenia Exome Sequencing Meta-Analysis (SCHEMA) consortium have identified that protein-truncating variants of exportin 7 (XPO7) can increase the risk of schizophrenia (odds ratio, 28.1). Here we show that mice with Xpo7 haploinsufficiency (Xpo7^+/- mice) present with cognitive and social behavioral impairments. Through proteome analysis using immunoprecipitation and frontal cortex nuclear isolation of Xpo7^+/- mice, we identify 45 molecules interacting with Xpo7, including CutC, Rbfox3, and Gria3. Through single-nucleus RNA sequencing of the frontal cortex and striatum of Xpo7^+/- mice differentiating between the onset and progressive stages, we also identify 284 gene expression changes that correlate with these stages. These genes encompass high-odds risk genes of schizophrenia identified by SCHEMA, including Gria3, Grin2A, Herc1, and Trio. Furthermore, our approach reveals 15 gene expression changes in the frontal cortex that correlate with the progressive stages. Our findings indicate the importance of investigating whether the interactions among the high-risk genes identified by SCHEMA contribute to a common schizophrenia pathology and underscore the significance of stage-dependent analysis.

Although females typically have a survival advantage, those with PTEN functional abnormalities face a higher risk of developing tumors than males. However, the differences in how each sex responds to PTEN dysfunction have rarely been studied. We use Caenorhabditis elegans to investigate how male and hermaphrodite worms respond to dysfunction of the PTEN homolog daf-18. Our study reveals that male worms can counterbalance the negative effects of daf-18 deficiency, resulting in longer adult lifespan. The survival advantage depends on the loss of DAF-18 protein phosphatase activity, while its lipid phosphatase activity is dispensable. The deficiency in DAF-18 protein phosphatase activity leads to the failure of dephosphorylation of the endoplasmic reticulum membrane protein C18E9.2/SEC62, causing increased levels of unfolded and aggregated proteins in hermaphrodites. In contrast, males maintain proteostasis through a UNC-23/NEF-mediated protein ubiquitination and degradation process, providing them with a survival advantage. We find that sex determination is a key factor in regulating the differential expression of unc-23 between sexes in response to daf-18 loss. These findings highlight the unique role of the male sex determination pathway in regulating protein degradation.

Immune checkpoint inhibitors against PD-1/PD-L1 are highly effective in immunologically hot tumours such as triple-negative breast cancer, wherein constitutive DNA damage promotes inflammation, while inducing PD-L1 expression to avoid attack by cytotoxic T cells. However, whether and how PD-L1 regulates the DNA damage response and inflammation remains unclear. Here, we show that nuclear PD-L1 activates the ATR-Chk1 pathway and induces proinflammatory chemocytokines upon genotoxic stress. PD-L1 interacts with ATR and is essential for Chk1 activation and chromatin binding. cGAS-STING and NF-κB activation in the late phase of the DNA damage response is inhibited by PD-L1 deletion or by inhibitors of ATR and Chk1. Consequently, the induction of proinflammatory chemocytokines at this stage is inhibited by deletion of PD-L1, but restored by the ATR activator Garcinone C. Inhibition of nuclear localisation by PD-L1 mutations or the HDAC2 inhibitor Santacruzamate A inhibits chemocytokine induction. Conversely, the p300 inhibitor C646, which accelerates PD-L1 nuclear localisation, promotes chemocytokine induction. These findings suggest that nuclear PD-L1 strengthens the properties of hot tumours and contributes to shaping the tumour microenvironment.

The protein interactome of p65/RELA, the most active subunit of the transcription factor (TF) NF-κB, has not been previously determined in living cells. Using p65-miniTurbo fusion proteins and biotin tagging, we identify >350 RELA interactors from untreated and IL-1α-stimulated cells, including many TFs (47% of all interactors) and >50 epigenetic regulators belonging to different classes of chromatin remodeling complexes. A comparison with the interactomes of two point mutants of p65 reveals that the interactions primarily require intact dimerization rather than DNA-binding properties. A targeted RNAi screen for 38 interactors and subsequent functional transcriptome and bioinformatics studies identify gene regulatory (sub)networks, each controlled by RELA in combination with one of the TFs ZBTB5, GLIS2, TFE3/TFEB, or S100A8/A9. The large, dynamic and versatile high-resolution interactome of RELA and its gene regulatory logics provides a rich resource and a new framework for explaining how RELA cooperativity determines gene expression patterns.

Live imaging of secretory cargoes is a powerful method for understanding the mechanisms of membrane trafficking. Inducing the synchronous release of cargoes from an organelle is key for enhancing microscopic observation. We developed an optical cargo-releasing method, 'retention using dark state of LOV2' (RudLOV), which enables precise spatial, temporal, and quantity control during cargo release. A limited amount of cargo-release using RudLOV is able to visualize cargo cisternal-movement and cargo-specific exit sites on the Golgi/trans-Golgi network. Moreover, by controlling the timing of cargo-release using RudLOV, we reveal the canonical and non-canonical effects of the well-known dynamin inhibitor dynasore, which inhibits early- but not late-Golgi transport and exits from the trans-Golgi network where dynamin-2 is active. Accumulation of COPI vesicles at the cis-side of the Golgi stacks in dynasore-treated cells suggests that dynasore targets COPI-uncoating/tethering/fusion machinery in the early-Golgi cisternae or endoplasmic reticulum but not in the late-Golgi cisternae. These results provide insight into the cisternal maturation of Golgi stacks.

Viral infection activates the transcription factors IRF3 and NF-κB, which induce type I interferon (IFN) and antiviral innate immune responses. Here, we identify dual-specific tyrosine phosphorylation-regulated kinase 4 (DYRK4) as an important regulator of virus-triggered IFN-β induction and antiviral innate immunity. Overexpression of DYRK4 enhances virus-triggered activation of IRF3 and type I IFN induction, whereas knockdown or knockout of DYRK4 impairs virus-induced activation of IRF3 and NF-κB. Moreover, Dyrk4-knockout mice are more susceptible to viral infection. The underlying mechanism involves DYRK4 acting as a scaffold protein to recruit TRIM71 and LUBAC to IRF3, increasing IRF3 linear ubiquitination, maintaining IRF3 stability and activation during viral infection, and promoting the IRF3-mediated antiviral response. Our findings provide new insights into the molecular mechanisms underlying viral infection-triggered IRF3 stabilization and activation.

microRNAs (miRNAs) are important post-transcriptional regulators that activate silencing mechanisms by annealing to mRNA transcripts. While plant miRNAs match their targets with nearly-full complementarity leading to mRNA cleavage, miRNAs in most animals require only a short sequence called 'seed' to inhibit target translation. Recent findings showed that miRNAs in cnidarians, early-branching metazoans, act similarly to plant miRNAs, by exhibiting full complementarity and target cleavage; however, it remained unknown if seed-based regulation was possible in cnidarians. Here, we investigate the miRNA-target complementarity requirements for miRNA activity in the cnidarian Nematostella vectensis. We show that bilaterian-like complementarity of seed-only or seed and supplementary 3' matches are insufficient for miRNA-mediated knockdown. Furthermore, miRNA-target mismatches in the cleavage site decrease knockdown efficiency. Finally, miRNA silencing of a target with three seed binding sites in the 3' untranslated region that mimics typical miRNA targeting was repressed in zebrafish but not in Nematostella and another cnidarian, Hydractinia symbiolongicarpus. Altogether, these results unravel striking similarities between plant and cnidarian miRNAs supporting a possible common evolutionary origin of miRNAs in plants and animals.

Ductular reaction (DR) is the hallmark of cholestatic diseases manifested in the proliferation of bile ductules lined by biliary epithelial cells (BECs). It is commonly associated with an increased risk of fibrosis and liver failure. The receptor for advanced glycation end products (RAGE) was identified as a critical mediator of DR during chronic injury. Yet, the direct link between RAGE-mediated DR and fibrosis as well as the mode of interaction between BECs and hepatic stellate cells (HSCs) to drive fibrosis remain elusive. Here, we delineate the specific function of RAGE on BECs during DR and its potential association with fibrosis in the context of cholestasis. Employing a biliary lineage tracing cholestatic liver injury mouse model, combined with whole transcriptome sequencing and in vitro analyses, we reveal a role for BEC-specific Rage activity in fostering a pro-fibrotic milieu. RAGE is predominantly expressed in BECs and contributes to DR. Notch ligand Jagged1 is secreted from activated BECs in a Rage-dependent manner and signals HSCs in trans, eventually enhancing fibrosis during cholestasis.