Targeting RNA G-quadruplex with repurposed drugs blocks SARS-CoV-2 entry.

The rapid emergence of SARS-CoV-2 variants of concern, the complexity of infection, and the functional redundancy of host factors, underscore an urgent need for broad-spectrum antivirals against the continuous COVID-19 pandemic, with drug repurposing as a viable therapeutic strategy. Here we report the potential of RNA G-quadruplex (RG4)-targeting therapeutic strategy for SARS-CoV-2 entry. Combining bioinformatics, biochemical and biophysical approaches, we characterize the existence of RG4s in several SARS-CoV-2 host factors. In silico screening followed by experimental validation identify Topotecan (TPT) and Berbamine (BBM), two clinical approved drugs, as RG4-stabilizing agents with repurposing potential for COVID-19. Both TPT and BBM can reduce the protein level of RG4-containing host factors, including ACE2, AXL, FURIN, and TMPRSS2. Intriguingly, TPT and BBM block SARS-CoV-2 pseudovirus entry into target cells in vitro and murine tissues in vivo. These findings emphasize the significance of RG4 in SARS-CoV-2 pathogenesis and provide a potential broad-spectrum antiviral strategy for COVID-19 prevention and treatment.

A microRNA checkpoint for Ca2+ signaling and overload in acute pancreatitis.

Acute pancreatitis (AP) is a common digestive disease without specific treatment, and its pathogenesis features multiple deleterious amplification loops dependent on translation, triggered by cytosolic Ca2+ ([Ca2+]i) overload; however, the underlying mechanisms in Ca2+ overload of AP remains incompletely understood. Here we show that microRNA-26a (miR-26a) inhibits pancreatic acinar cell (PAC) store-operated Ca2+ entry (SOCE) channel expression, Ca2+ overload, and AP. We find that major SOCE channels are post-transcriptionally induced in PACs during AP, whereas miR-26a expression is reduced in experimental and human AP and correlated with AP severity. Mechanistically, miR-26a simultaneously targets Trpc3 and Trpc6 SOCE channels and attenuates physiological oscillations and pathological elevations of [Ca2+]i in PACs. MiR-26a deficiency increases SOCE channel expression and [Ca2+]i overload, and significantly exacerbates AP. Conversely, global or PAC-specific overexpression of miR-26a in mice ameliorates pancreatic edema, neutrophil infiltration, acinar necrosis, and systemic inflammation, accompanied with remarkable improvements on pathological determinants related with [Ca2+]i overload. Moreover, pancreatic or systemic administration of an miR-26a mimic to mice significantly alleviates experimental AP. These findings reveal a previously unknown mechanism underlying AP pathogenesis, establish a critical role for miR-26a in Ca2+ signaling in the exocrine pancreas, and identify a potential target for the treatment of AP.

RNA G-quadruplex in TMPRSS2 reduces SARS-CoV-2 infection.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection continues to have devastating consequences worldwide. Recently, great efforts have been made to identify SARS-CoV-2 host factors, but the regulatory mechanisms of these host molecules, as well as the virus per se, remain elusive. Here we report a role of RNA G-quadruplex (RG4) in SARS-CoV-2 infection. Combining bioinformatics, biochemical and biophysical assays, we demonstrate the presence of RG4s in both SARS-CoV-2 genome and host factors. The biological and pathological importance of these RG4s is then exemplified by a canonical 3-quartet RG4 within Tmprss2, which can inhibit Tmprss2 translation and prevent SARS-CoV-2 entry. Intriguingly, G-quadruplex (G4)-specific stabilizers attenuate SARS-CoV-2 infection in pseudovirus cell systems and mouse models. Consistently, the protein level of TMPRSS2 is increased in lungs of COVID-19 patients. Our findings reveal a previously unknown mechanism underlying SARS-CoV-2 infection and suggest RG4 as a potential target for COVID-19 prevention and treatment.

UBE2O promotes lipid metabolic reprogramming and liver cancer progression by mediating HADHA ubiquitination.

Cancer cells rely on heightened protein quality control mechanisms, including the ubiquitin-proteosome system that is predominantly driven by ubiquitination comprising E1, E2, and E3 trienzyme cascades. Although E3s have been extensively studied, the implication of E2s in tumorigenesis is poorly defined. Here we reveal a critical E2 in the pathogenesis of hepatocellular carcinoma (HCC). Among all of E2s, UBE2O shows the strongest association with HCC survival prognosis, and its expression is increased in HCC tumors. Accordingly, UBE2O deficiency inhibits HCC growth and metastasis both in vitro and in vivo, while its overexpression has opposite effects. Depending on both E2 and E3 enzymatic activities, UBE2O can interact with and mediate the ubiquitination and degradation of HADHA, a mitochondrial ß-oxidation enzyme, thereby modulating lipid metabolic reprogramming. HADHA is reduced in HCC tumors and inversely correlated with UBE2O levels. Importantly, HADHA acts as a tumor suppressor and primarily mediates UBE2O's function on HCC. Moreover, liver-specific deletion of Ube2o in mice are resistant to DEN-induced hepatocarcinogenesis, along with HADHA upregulation and reduced hepatic lipid accumulation. These data reveal UBE2O as a novel oncogenic driver for metabolic reprogramming and HCC development, highlighting the potential of targeting UBE2O/HADHA axis for HCC therapy.

Pyroptosis of MCF7 Cells Induced by the Secreted Factors of hUCMSCs.

Human umbilical cord mesenchymal stem cells (hUCMSCs) are superior to other sources of mesenchymal stem/stromal cells (MSCs), and they are used as a novel tool for cell-based cancer therapy. However, the mechanism underlying hUCMSC-induced cancer cell death is not clear. In the present study, we aimed to evaluate the effect of secreted factors of hUCMSCs on the breast cancer cell line MCF7 by exposing them to the conditioned medium (CM) of hUCMSCs. We evaluated the morphological changes, cell viability, cell cycle, apoptosis, DNA fragmentation, and interleukin-1ß (IL-1ß) secretion of CM-exposed MCF7 cells. The results showed that the secreted factors of hUCMSCs could cause MCF7 cell death by inducing pyroptosis. We also sequenced the total RNA, and the differentially expressed genes (DEGs) were subjected to the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis. A total of 2597 (1822 upregulated and 775 downregulated) genes were identified and 14 pathways were significantly enriched. The results showed that the expression of the pyroptosis-related genes NLRP1 and CASP4 and the inflammation-related pathways changed significantly in MCF7 cells exposed to the CM. To the best of our knowledge, this study is the first to report that the secreted factors of hUCMSCs can cause MCF7 cell pyroptosis. Furthermore, it is the first to examine the global gene expression in MCF7 cells exposed to CM. These results will provide valuable information for further studies on the mechanism of MCF7 cell pyroptosis induced by the secreted factors of hUCMSCs. It will also help understand the effect of hUCMSCs on cell-based breast cancer therapy.

Heterogenic transplantation of bone marrow-derived rhesus macaque mesenchymal stem cells ameliorates liver fibrosis induced by carbon tetrachloride in mouse.

Liver fibrosis is a disease that causes high morbidity and has become a major health problem. Liver fibrosis can lead to the end stage of liver diseases (livercirrhosisand hepatocellularcarcinoma). Currently, liver transplantation is the only effective treatment for end-stage liver disease. However, the shortage of organ donors, high cost of medical surgery, immunological rejection and transplantation complications severely hamper liver transplantation therapy. Mesenchymal stem cells (MSCs) have been regarded as promising cells for clinical applications in stem cell therapy in the treatment of liver diseases due to their unique multipotent differentiation capacity, immunoregulation and paracrine effects. Although liver fibrosis improvements by MSC transplantation in preclinical experiments as well as clinical trials have been reported, the in vivo fate of MSCs after transportation and their therapeutic mechanisms remain unclear. In this present study, we isolated MSCs from the bone marrow of rhesus macaques. The cells exhibited typical MSC markers and could differentiate into chondrocytes, osteocytes, and adipocytes, which were not affected by labeling with enhanced green fluorescent protein (EGFP). The harvested MSCs respond to interferon-γ stimulation and have the ability to inhibit lymphocyte proliferation in vitro. EGFP-labeled MSCs (1 × 106 cells) were transplanted into mice with carbon tetrachloride-induced liver fibrosis via tail vein injection. The ability of the heterogenic MSC infusion to ameliorate liver fibrosis in mice was evaluated by a blood plasma chemistry index, pathological examination and liver fibrosis-associated gene expression. Additionally, a small number of MSCs that homed and engrafted in the mouse liver tissues were evaluated by immunofluorescence analysis. Our results showed that the transplantation of heterogenic MSCs derived from monkey bone marrow can be used to treat liver fibrosis in the mouse model and that the paracrine effects of MSCs may play an important role in the improvement of liver fibrosis.