摘要：MicroRNAs (miRNAs) have been demonstrated for their involvement in virus biology and pathogenesis, including functioning as key determinants of virally-induced cancers. As an important oncogenic α-herpesvirus affecting poultry health, Marek’s disease virus serotype 1 [Gallid alphaherpesvirus2 (GaHV-2)] induces rapid-onset T-cell lymphomatous disease commonly referred to as Marek’s disease (MD), an excellent biological model for the study of virally-induced cancer in the natural hosts. Previously, we have demonstrated that GaHV-2-encoded miRNAs (especially those within the Meq-cluster) have the potential to act as critical regulators of multiple processes such as virus replication, latency, pathogenesis, and/or oncogenesis. In addition to miR-M4-5p (miR-155 homolog) and miR-M3-5p, we have recently found that miR-M2-5p possibly participate in inducing MD lymphomagenesis. Here, we report the identification of two tumor suppressors, the RNA-binding protein 24 (RBM24) and myogenic differentiation 1 (MYOD1), being two biological targets for miR-M2-5p. Our experiments revealed that as a critical miRNA, miR-M2-5p promotes cell proliferationviaregulating the RBM24-mediated p63 overexpression and MYOD1-mediated IGF2 signaling and suppresses apoptosis by targeting the MYOD1-mediated Caspase-3 signaling pathway. Our data present a new strategy of a single viral miRNA exerting dual role to potentially participate in the virally-induced T-cell lymphomagenesis by simultaneously promoting the cell proliferation and suppressing apoptosis.

摘要：Many rice microRNAs have been identified as fine-tuning factors in the regulation of agronomic traits and immunity. Among them, Osa-miR535 targetsSQUAMOSA promoter binding protein-like 14(OsSPL14) to positively regulate tillers but negatively regulate yield and immunity. Here, we uncovered that Osa-miR535 targets anotherSPLgene,OsSPL4, to suppress rice immunity againstMagnaporthe oryzae. Overexpression of Osa-miR535 significantly decreased the accumulation of the fusion protein SPL4TBS-YFP that contains the target site of Osa-miR535 inOsSPL4. Consistently, Osa-miR535 mediated the cleavage ofOsSPL4mRNA between the 10th and 11th base pair of the predicted binding site at the 3′ untranslated region. Transgenic rice lines overexpressingOsSPL4(OXSPL4) displayed enhanced blast disease resistance accompanied by enhanced immune responses, including increased expression of defense-relative genes and up-accumulated H2O2. By contrast, the knockout mutantosspl4exhibited susceptibility. Moreover, OsSPL4 binds to the promoter ofGH3.2, an indole-3-acetic acid-amido synthetase, and promotes its expression. Together, these data indicate thatOs-miR535 targetsOsSPL4andOsSPL4-GH3.2, which may parallel theOsSPL14-WRKY45module in rice blast disease resistance.

摘要：Employing two different alkyne-modified dopamine agonists to construct bivalent compounds via click chemistry resulted in the identification of a bivalent ligand (11c) for dopamine D2 receptor homodimer, which, compared to its parent monomeric alkyne, showed a 16-fold higher binding affinity for the dopamine D2 receptor and a 5-fold higher potency in a cAMP assay in HEK 293T cells stably expressing D2R. Molecular modeling revealed that 11c can indeed bridge the orthosteric binding sites of a D2R homodimer in a relaxed conformation via the TM5-TM6 interface and allows to largely rationalize the results of the receptor assays.

摘要：Most persimmon (Diospyros kaki) cultivars are astringent and require post-harvest deastringency treatments such as 95% CO2(high-CO2treatment) to make them acceptable to consumers. High-CO2treatment can, however, also induce excessive softening, which can be reduced by adding 1-methylcyclopropene (1-MCP). Previous studies have shown that genes encoding the ETHYLENE RESPONSE FACTORS (ERFs) DkERF8/16/19 can trans-activatexyloglucan endotransglycosylase/hydrolase(DkXTH9), which encodes the cell wall-degrading enzyme associated with persimmon fruit softening. In this study, RNA-seq data between three treatments were compared, namely high-CO2, high-CO2+1-MCP, and controls. A total of 227 differentially expressed genes, including 17 transcription factors, were predicted to be related to persimmon post-deastringency softening. Dual-luciferase assays indicated that DkNAC9 activated theDkEGase1promoter 2.64-fold. Synergistic effects on transcription ofDkEGase1that involved DkNAC9 and the previously reported DkERF8/16 were identified. Electrophoretic mobility shift assay indicated that DkNAC9 could physically bind to theDkEGase1promoter. Bimolecular fluorescence complementation and firefly luciferase complementation imaging assays indicated protein–protein interactions between DkNAC9 and DkERF8/16. Based on these findings, we conclude that DkNAC9 is a direct transcriptional activator ofDkEGase1that can co-operate with DkERF8/16 to enhance fruit post-deastringency softening.

摘要：The fungusNosema bombyciscauses significant economic losses via parasitism of an economically important insect. MicroRNAs (miRNAs) play important roles in regulating host and parasite gene expression via mRNA degradation or by inhibiting protein translation. To investigate whether microRNA-like RNAs (milRNAs) regulateN. bombycispathogenesis and to better understand the regulatory mechanisms underlying infection, we constructed small RNA libraries fromN. bombycishyphae during the schizont proliferation period. Eleven novel milRNAs were determined by RNA sequencing and stem-loop reverse transcriptase PCR (RT-PCR) assays. Moreover, a virulence-associated milRNA, Nb-milR8, was identified as critical forN. bombycisproliferation by binding and downregulating expression of its target gene,BmPEX16, in the host during infection. Silencing of Nb-milR8 or overexpression of the targetBmPEX16gene resulted in increased susceptibility ofBombyx moritoN. bombycisinfection. Taken together, these results suggest that Nb-milR8 is an important virulence factor that acts as an effector to suppress host peroxidase metabolism, thereby facilitatingN. bombycisproliferation. These results provide important novel insights into interactions between pathogenic fungi and their hosts.

摘要：Nucleotide-binding leucine-rich repeat (NLR) proteins work as crucial intracellular immune receptors. N-terminal domains of NLRs fall into two groups, coiled-coil (CC) and Toll-interleukin 1 receptor domains, which play critical roles in signal transduction and disease resistance. However, the activation mechanisms of NLRs, and how their N-termini function in immune induction, remain largely unknown. Here, we revealed that the CC domain of a rice NLR Pit contributes to self-association. The Pit CC domain possesses three conserved hydrophobic residues that are known to be involved in oligomer formation in two NLRs, barley MLA10 andArabidopsisRPM1. Interestingly, the function of these residues in Pit differs from that in MLA10 and RPM1. Although three hydrophobic residues are important for Pit-induced disease resistance against rice blast fungus, they do not participate in self-association or binding to downstream signalling molecules. By homology modelling of Pit using the Arabidopsis ZAR1 structure, we tried to clarify the role of three conserved hydrophobic residues and found that they are located in the predicted α2-helix of the Pit CC domain and involved in the plasma membrane localization. Our findings provide novel insights for understanding the mechanisms of NLR activation as well as the relationship between subcellular localization and immune induction.

摘要：Theaquatic environmentcan be greatly impacted by thermal and hypoxic stresses, particularly caused by intensified global warming. Hence, there is an urgency to understand the response mechanisms of marine organisms to adverse environment. Although long non-coding RNAs (lncRNAs) are involved in many biological processes, their roles in stress responses still remain unclear. Here, differentially expressed (DE) lncRNAs and mRNAs were identified as responses to environmental stresses in the economically important sea cucumber,Apostichopus japonicus, and their potential roles were explored. Based on a total of 159, 355 and 495 significantly upregulated genes and 230, 518 and 647 significantly downregulated genes identified in the thermal, hypoxic and combination thermal + hypoxic stress treatments, respectively, we constructed DE-lncRNA-mRNA coexpression networks. Among the networks, eight shared pairs were identified from the three treatments, and based on the connectivity degree, MSTRG.27265, MSTRG.19729 and MSTRG.95524 were shown to be crucial lncRNAs. Among all the significantly changed lncRNAs identified by RT-qPCR and sequencing data, binding sites were found in four other lncRNAs (MSTRG.34610, MSTRG.10941, MSTRG.81281 and MSTRG.93731) with Aja-miR-2013-3p, a key miRNA that responds to hypoxia in sea cucumbers. The hypoxia-inducible factor (HIF-1α) was also shown as the possible targetedmRNAof Aja-miR-2013-3p. As indicated by a dual-luciferase reporter assay system, “HIF-1α gene/Aja-miR-2013-3p/MSTRG.34610” network and the “HIF-1α gene/Aja-miR-2013-3p/MSTRG.10941” network may play important roles in sea cucumbers under environmental stresses. Moreover, environmental stress altered the expression of multiple lncRNAs and mRNAs, thus affecting various biological processes inA. japonicus, including immunity, energy metabolism and the cell cycle. At the molecular level, more comprehensive responses were elicited by the combined thermal/hypoxic stress treatment than by individual stresses alone in sea cucumbers. This study lays the groundwork for future research on molecular mechanisms ofechinodermresponses to thermal and hypoxic stress in the context of global climate changes.

摘要：Aberrant activation of NLRP3 inflammasome has been implicated in a variety of human inflammatory diseases, but currently, no pharmacological NLRP3 inhibitor has been approved. In this study, we showed that echinatin, the ingredient of the traditional herbal medicine licorice, effectively suppresses the activation of NLRP3 inflammasome in vitro and in vivo. Further investigation revealed that echinatin exerts its inhibitory effect on NLRP3 inflammasome by binding to heat-shock protein 90 (HSP90), inhibiting its ATPase activity and disrupting the association between the cochaperone SGT1 and HSP90-NLRP3. Importantly, in vivo experiments demonstrated that administration of echinatin obviously inhibits NLRP3 inflammasome activation and ameliorates LPS-induced septic shock and dextran sodium sulfate–induced (DSS-induced) colitis in mice. Moreover, echinatin exerted favorable pharmacological effects on liver inflammation and fibrosis in a mouse model of nonalcoholic steatohepatitis (NASH). Collectively, our study identifies echinatin as a potentially novel inhibitor of NLRP3 inflammasome, and its use may be developed as a therapeutic approach for the treatment of NLRP3-driven diseases.

摘要：Oesophageal squamous cell carcinoma (ESCC) is a common and aggressive malignancy. Although many molecular alterations have been observed in ESCC, the mechanisms underlying the development and progression of this disease remain unclear. In the present study, miR-1224-5p was identified to be downregulated in ESCC tissues compared to normal tissues, and its low expression was correlated with shorter survival time in patients. In vitro experiments showed that miR-1224-5p inhibited the proliferation, colony formation, migration and invasion of ESCC cells. Mechanistic investigation revealed that miR-1224-5p directly targeted TNS4 and inhibited its expression, which led to the inactivation of EGFR-EFNA1/EPHA2-VEGFA (vascular endothelial growth factor A) signalling. Experiments in vivo confirmed the suppressive effect of miR-1224-5p on oesophageal cancer cells. By immunohistochemistry analysis of ESCC specimens, we found that TNS4 expression was positively correlated with that of VEGFA, and was significantly associated with lymph node metastasis and shorter survival time in patients. Together, our data suggest that miR-1224-5p downregulation is a frequent alteration in ESCC that promotes cell proliferation, migration, invasion and tumour growth by activating the EGFR-EFNA1/EPHA2-VEGFA signalling pathway via inhibition of TNS4 expression. Decreased miR-1224-5p and elevated TNS4 are unfavourable prognostic factors for ESCC patients.

摘要：Accumulating evidence indicates that cryptochrome circadian regulatory (CRY) proteins have emerged as crucial regulators of osteogenic differentiation. However, the associated mechanisms are quite elusive. In this study, we show that knockdown of CRY2 downregulated the expression of runt-related transcription factor 2 (Runx2), alkaline phosphatase (ALP), osteocalcin (OCN), and osteopontin (OPN) to facilitate osteoblast differentiation. Further study identified that CRY2 was directly targeted by microRNA (miR)-7-5p, which was highly induced during osteoblast differentiation. The expression of Runx2, ALP, collagen type I alpha 1 (Col1a1), and OCN was upregulated by overexpression of miR-7-5p and induction of osteoblast differentiation. Moreover, signal transducer and activator of transcription 3 (STAT3) transcriptionally activated miR-7-5p to significantly enhance the expression of above osteogenic marker genes and mineral formation. However, overexpression of CRY2 abolished the osteogenic differentiation induced by miR-7-5p overexpression. Silencing of CRY2 unraveled the binding of CRY2 with the circadian locomotor output cycles kaput (CLOCK)/brain and muscle ARNT-like 1 (BMAL1) complex to release CLOCK/BMAL1, which facilitated the binding of CLOCK/BMAL1 to the promoter region of the P300 E-box to stimulate the transcription of P300. P300 subsequently promoted the acetylation of histone 3 and the formation of a transcriptional complex with Runx2 to enhance osteogenesis. Taken together, our study revealed that CRY2 is repressed by STAT3/miR-7-5p to promote osteogenic differentiation through CLOCK/BMAL1/P300 signaling. The involved molecules may be potentially targeted for treatment of osteoporosis.