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Characterized by progressive degeneration of retinal ganglion cells (RGCs) and vision loss, glaucoma is the primary cause of irreversible blindness, incurable and affecting over 78 million patients. However, pathogenic mechanisms leading to glaucoma-induced RGC loss are incompletely understood. Unexpectedly, we found that cGAS-STING (2'3'-cyclic GMP-AMP-stimulator of interferon genes) signaling, which surveils displaced double-stranded DNA (dsDNA) in the cytosol and initiates innate immune responses, was robustly activated during glaucoma in retinal microglia in distinct murine models. Global or microglial deletion of STING markedly relieved glaucoma symptoms and protected RGC degeneration and vision loss, while mice bearing genetic cGAS-STING supersensitivity aggravated retinal neuroinflammation and RGC loss. Mechanistically, dsDNA from tissue injury activated microglial cGAS-STING signaling, causing deleterious macroglia reactivity in retinas by cytokine-mediated microglia-macroglia interactions, progressively driving apoptotic death of RGCs. Remarkably, preclinical investigations of targeting cGAS-STING signaling by intraocular injection of TBK1i or anti-IFNAR1 antibody prevented glaucoma-induced losses of RGCs and vision. Therefore, we unravel an essential role of cGAS-STING signaling underlying glaucoma pathogenesis and suggest promising therapeutic strategies for treating this devastating disease. [ABSTRACT FROM AUTHOR]

Cytosolic double-stranded DNA surveillance by cyclic GMP-AMP synthase (cGAS)-Stimulator of Interferon Genes (STING) signaling triggers cellular senescence, autophagy, biased mRNA translation, and interferon-mediated immune responses. However, detailed mechanisms and physiological relevance of STING-induced senescence are not fully understood. Here, we unexpectedly found that interferon regulatory factor 3 (IRF3), activated during innate DNA sensing, forms substantial endogenous complexes in the nucleus with retinoblastoma (RB), a key cell cycle regulator. The IRF3-RB interaction attenuates cyclin-dependent kinase 4/6 (CDK4/6)-mediated RB hyperphosphorylation that mobilizes RB to deactivate E2 family (E2F) transcription factors, thereby driving cells into senescence. STING-IRF3-RB signaling plays a notable role in hepatic stellate cells (HSCs) within various murine models, pushing activated HSCs toward senescence. Accordingly, IRF3 global knockout or conditional deletion in HSCs aggravated liver fibrosis, a process mitigated by the CDK4/6 inhibitor. These findings underscore a straightforward yet vital mechanism of cGAS-STING signaling in inducing cellular senescence and unveil its unexpected biology in limiting liver fibrosis. [ABSTRACT FROM AUTHOR]

Inhibition of heat shock protein 90 (Hsp90), a prominent molecular chaperone, effectively limits severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection but little is known about any interaction between Hsp90 and SARS-CoV-2 proteins. Here, we systematically analyzed the effects of the chaperone isoforms Hsp90a and Hsp90β on individual SARSCoV-2 viral proteins. Five SARS-CoV-2 proteins, namely nucleocapsid (N), membrane (M), and accessory proteins Orf3, Orf7a, and Orf7b were found to be novel clients of Hsp90β in particular. Pharmacological inhibition of Hsp90 with 17-DMAG results in N protein proteasome-dependent degradation. Hsp90 depletion-induced N protein degradation is independent of CHIP, a ubiquitin E3 ligase previously identified for Hsp90 client proteins, but alleviated by FBXO10, an E3 ligase identified by subsequent siRNA screening. We also provide evidence that Hsp90 depletion may suppress SARS-CoV-2 assembly partially through induced M or N degradation. Additionally, we found that GSDMD-mediated pyroptotic cell death triggered by SARSCoV-2 was mitigated by inhibition of Hsp90. These findings collectively highlight a beneficial role for targeting of Hsp90 during SARS-CoV-2 infection, directly inhibiting virion production and reducing inflammatory injury by preventing the pyroptosis that contributes to severe SARS-CoV-2 disease. [ABSTRACT FROM AUTHOR]

Bone morphogenetic proteins (BMPs) play vital roles in regulating stem cell maintenance and differentiation. BMPs can induce osteogenesis and inhibit myogenesis of mesenchymal stem cells. Canonical BMP signaling is stringently controlled through reversible phosphorylation and nucleocytoplasmic shuttling of Smad1, Smad5, and Smad8 (Smad1/5/8). However, how the nuclear export of Smad1/5/8 is regulated remains unclear. Here we report that the Ran-binding protein RanBP3L acts as a nuclear export factor for Smad1/5/8. RanBP3L directly recognizes dephosphorylated Smad1/5/8 and mediates their nuclear export in a Randependent manner. Increased expression of RanBP3L blocks BMP-induced osteogenesis of mouse bone marrow-derived mesenchymal stem cells and promotes myogenic induction of C2C12 mouse myoblasts, whereas depletion of RanBP3L expression enhances BMP-dependent stem cell differentiation activity and transcriptional responses. In conclusion, our results demonstrate that RanBP3L, as a nuclear exporter for BMP-specific Smads, plays a critical role in terminating BMP signaling and regulating mesenchymal stem cell differentiation. [ABSTRACT FROM AUTHOR]

Adherens junction remodeling regulated by apical polarity proteins constitutes a major driving force for tissue morphogenesis, although the precise mechanism remains inconclusive. Here, we report that, in zebrafish, the Crumbs complex component MPP5a interacts with small GTPase Rab11 in Golgi to transport cadherin and Crumbs components synergistically to the apical domain, thus establishing apical epithelial polarity and adherens junctions. In contrast, Par complex recruited byMPP5a is incapable of interacting with Rab11 but might assemble cytoskeleton to facilitate cadherin exocytosis. In accordance, dysfunction of MPP5a induces an invasive migration of epithelial cells. This adherens junction remodeling pattern is frequently observed in zebrafish lens epithelial cells and neuroepithelial cells. The data identify an unrecognized MPP5a-Rab11 complex and describe its essential role in guiding apical polarization and zonula adherens formation in epithelial cells. [ABSTRACT FROM AUTHOR]

A proper inflammatory response is critical to the restoration of tissue homeostasis after injury or infection, but how such a response is modulated by the physical properties of the cellular and tissue microenvironments is not fully understood. Here, using H358, HeLa, and HEK293T cells, we report that cell density can modulate inflammatory responses through the Hippo signaling pathway.Wefound that NF-ĸB activation through the proinflammatory cytokines interleukin-1β (IL-1β) and tumor necrosis factor α (TNFα) is not affected by cell density. However, we also noted that specific NF-ĸB target genes, such as cyclooxygenase 2 (COX-2), are induced much less at low cell densities than at high cell densities. Mechanistically, we observed that the transcriptional coactivators Yes-associated protein (YAP) and transcriptional coactivator with PDZ-binding motif (TAZ) are localized to the nucleus, bind to TEA domain transcription factors (TEADs), recruit histone deacetylase 7 (HDAC7) to the promoter region of COX-2, and repress its transcription at low cell density and that high cell density abrogates this YAP/TAZ-mediated transcriptional repression. Of note, IL-1β stimulation promoted cell migration and invasion mainly through COX-2 induction, but YAP inhibited this induction and thus cell migration and invasion. These results suggest that YAP/TAZ--TEAD interactions can repress COX-2 transcription and thereby mediate cell density--dependent modulation of proinflammatory responses. Our findings highlight that the cellular microenvironment significantly influences inflammatory responses via the Hippo pathway. [ABSTRACT FROM AUTHOR]

Smad7 is a negative feedback product of TGF-β superfamily signaling and fine tunes a plethora of pleiotropic responses induced by TGF-β ligands. However, its noncanonical functions independent of TGF-β signaling remain to be elucidated. Here, we show that Smad7 activates signal transducers and activators of transcription 3 (STAT3) signaling in maintaining mouse embryonic stem cell pluripotency in a manner independent of the TGF-β receptors, yet dependent on the leukemia inhibitory factor (LIF) coreceptor glycoprotein 130 (gp130). Smad7 directly binds to the intracellular domain of gp130 and disrupts the SHP2-gp130 or SOCS3-gp130 complex, thereby amplifying STAT3 activation. Consequently, Smad7 facilitates LIF-mediated self-renewal of mouse ESCs and is also critical for induced pluripotent stem cell reprogramming. This finding illustrates an uncovered role of the Smad7-STAT3 interplay in maintaining cell pluripotency and also implicates a mechanism involving Smad7 underlying cytokine-dependent regulation of cancer and inflammation. [ABSTRACT FROM AUTHOR]

The epithelial-to-mesenchymal transition (EMT) is a process by which differentiated epithelial cells reprogram gene expression, lose their junctions and polarity, reorganize their cytoskeleton, increase cell motility and assume a mesenchymal morphology. Despite the critical functions of the microtubule (MT) in cytoskeletal organization, how it participates in EMT induction and maintenance remains poorly understood. Here we report that acetylated α-tubulin, which plays an important role in microtubule (MT) stabilization and cell morphology, can serve as a novel regulator and marker of EMT. A high level of acetylated α-tubulin was correlated with epithelial morphology and it profoundly decreased during TGF-β-induced EMT. We found that TGF-β increased the activity of HDAC6, a major deacetylase of α-tubulin, without affecting its expression levels. Treatment with HDAC6 inhibitor tubacin or TGF-β type I receptor inhibitor SB431542 restored the level of acetylated α-tubulin and consequently blocked EMT. Our results demonstrate that acetylated α-tubulin can serve as a marker of EMT and that HDAC6 represents an important regulator during EMT process. [ABSTRACT FROM AUTHOR]