野生青果的防腐作用研究

本文对野生青果的药用成分进行了鉴定及制备,证明了该成分具有较强的抑菌作用,并同常用的合成防腐剂苯甲酸钠、山梨酸钾的防腐性能进行了比较。     

Site-specific Acetylation of the Proteasome Activator REGγ Directs Its Heptameric Structure and Functions

The proteasome activator REGγ has been reported to promote degradation of steroid receptor coactivator-3 and cyclin-dependent kinase inhibitors p21, p16, and p19 in a ubiquitin- and ATP-independent manner. A recent comparative analysis of REGγ expression in mouse and human tissues reveals a unique pattern of REGγ in specific cell types, suggesting undisclosed functions and biological importance of this molecule. Despite the emerging progress made in REGγ-related studies, how REGγ function is regulated remains to be explored. In this study, we report for the first time that REGγ can be acetylated mostly on its lysine 195 (Lys-195) residue by CREB binding protein (CBP), which can be reversed by sirtuin 1 (SIRT1) in mammalian cells. Site-directed mutagenesis abrogated acetylation at Lys-195 and significantly attenuated the capability of REGγ to degrade its target substrates, p21 and hepatitis C virus core protein. Mechanistically, acetylation at Lys-195 is important for the interactions between REGγ monomers and ultimately influences REGγ heptamerization. Biological analysis of cells containing REGγ-WT or REGγ-K195R mutant indicates an impact of acetylation on REGγ-mediated regulation of cell proliferation and cell cycle progression. These findings reveal a previously unknown mechanism in the regulation of REGγ assembly and activity, suggesting a potential venue for the intervention of the ubiquitin-independent REGγ proteasome activity.     

The coiled-coil domain of TRAF6 is essential for its auto-ubiquitination

Tumor necrosis factor receptor-associated factor 6 (TRAF6) is a crucial signaling transducer that regulates a diverse array of physiological processes, including adaptive immunity, innate immunity, and bone metabolism. Importantly, it is essential for activating NF-kappaB signaling pathway in response to interleukin-1 and Toll-like receptor ligands. Previously, we characterized TRAF6 to be a ubiquitin ligase. In combination with the ubiquitin conjugating enzyme complex Ubc13/Uev1A, TRAF6 could catalyze the formation on itself of unique Lys-63 linked polyubiquitin chain that positively regulated NF-kappaB signaling pathway. However, it remains unknown how this auto-ubiquitination process is regulated. In this study, we found that the coiled-coil domain of TRAF6 was essential for its auto-ubiquitination and activating NF-kappaB signaling pathway. This domain served not as the specific target where the polyubiquitin chain was linked, but as a specific bridge to recruit Ubc13/Uev1A.     

Hsp90 regulates activation of interferon regulatory factor 3 and TBK-1 stabilization in Sendai virus-infected cells

Interferon regulatory factor 3 (IRF3) plays a crucial role in mediating cellular responses to virus intrusion. The protein kinase TBK1 is a key regulator inducing phosphorylation of IRF3. The regulatory mechanisms during IRF3 activation remain poorly characterized. In the present study, we have identified by yeast two-hybrid approach a specific interaction between IRF3 and chaperone heat-shock protein of 90 kDa (Hsp90). The C-terminal truncation mutant of Hsp90 is a strong dominant-negative inhibitor of IRF3 activation. Knockdown of endogenous Hsp90 by RNA interference attenuates IRF3 activation and its target gene expressions. Alternatively, Hsp90-specific inhibitor geldanamycin (GA) dramatically reduces expression of IRF3-regulated interferon-stimulated genes and abolishes the cytoplasm-to-nucleus translocation and DNA binding activity of IRF3 in Sendai virus-infected cells. Significantly, virus-induced IRF3 phosphorylation is blocked by GA, whereas GA does not affect the protein level of IRF3. In addition, TBK1 is found to be a client protein of Hsp90 in vivo. Treatment of 293 cells with GA interferes with the interaction of TBK1 and Hsp90, resulting in TBK1 destabilization and its subsequent proteasome-mediated degradation. Besides maintaining stability of TBK1, Hsp90 also forms a novel complex with TBK1 and IRF3, which brings TBK1 and IRF3 dynamically into proximity and facilitates signal transduction from TBK1 to IRF3. Our study uncovers an essential role of Hsp90 in the virus-induced activation of IRF3.     

MicroRNA-135b-5p Downregulation Causes Antidepressant Effects by Regulating SIRT1 Expression

Depression is a serious and potentially life-threatening mental illness. Recently, the role of sirtuin 1 (SIRT1) in chronic unpredictable mild stress (CUMS) management has been examined. The present study explored and clarified whether microRNA (miR)-135b-5p could play a role in depression by regulating the expression of SIRT1. SIRT1 was identified as the target gene of miR-135b-5p using TargetScan and the dual luciferase reporter assay. In addition, the expression levels of SIRT1 were significantly reduced in mouse peripheral blood and hippocampal tissue samples, while the expression of miR-135b-5p exhibited the opposite effects. Subsequently, the effects of miR-135b-5p inhibition were investigated in mice with depression. The results indicated that the miR-135b-5p inhibitor significantly increased the weight loss induced by CUMS compared with the model group, while reducing the expression levels of miR-135b-5p and further alleviating the depression-like behavior induced by CUMS. Concomitantly, the results indicated that the miR-135b-5p inhibitor inhibited CUMS-induced hippocampal cell apoptosis and significantly reduced the expression levels of cleaved caspase-3 and the ratio of cleaved caspase-3/caspase-3. Moreover, the miR-135b-5p inhibitor significantly reduced the CUMS-induced increase of the inflammatory factors IL-1β, IL-6 and TNF-α in the hippocampal mouse samples, while significantly increasing the expression levels of SIRT1. Finally, the results demonstrated that all the effects of the miR-135b-5p inhibitor on CUMS-induced mice were significantly reversed by SIRT1 silencing. In conclusion, the present study indicated that the miR-135b-5p/SIRT1 pathway was a key mediator of antidepressant effects induced in depressed mice. Therefore, it could be considered a potential therapeutic target for the treatment of CUMS-induced depression.

     

Herpes virus proteins ICP0 and BICP0 can activate NF-kappaB by catalyzing IkappaBalpha ubiquitination

The immediate early proteins ICP0 and BICP0 from Herpes virus are promiscuous activators of both viral and cellular genes and play a critical role in virus life cycle. Here we report that ICP0 and BICP0 could induce NF-kappaB translocation from cytoplasm into nucleus and strongly activate NF-kappaB responsive genes specifically. This process was dependent on the RING domain of both proteins. In addition, ICP0 interacted specifically with IkappaBalpha and its activating effect was attenuated by Ubch5A(C85A) and MG132, but not by IkappaBalpha(S32A/S36A). Remarkably, IkappaBalpha was poly-ubiquitinated by both ICP0 and BICP0, in vitro and in vivo. These data indicate that ICP0 and BICP0, functioning as ubiquitin ligases, are bona fide activators of NF-kappaB signaling pathway. Our study identifies a new way ICP0 and BICP0 explore to regulate gene expression.     

Regulation of REGγ cellular distribution and function by SUMO modification

Discovery of emerging REGγ-regulated proteins has accentuated the REGγ-proteasome as an important pathway in multiple biological processes, including cell growth, cell cycle regulation, and apoptosis. However, little is known about the regulation of the REGγ-proteasome pathway. Here we demonstrate that REGγ can be SUMOylated in vitro and in vivo by SUMO-1, SUMO-2, and SUMO-3. The SUMO-E3 protein inhibitor of activated STAT (PIAS)1 physically associates with REGγ and promotes SUMOylation of REGγ. SUMOylation of REGγ was found to occur at multiple sites, including K6, K14, and K12. Mutation analysis indicated that these SUMO sites simultaneously contributed to the SUMOylation status of REGγ in cells. Posttranslational modification of REGγ by SUMO conjugation was revealed to mediate cytosolic translocation of REGγ and to cause increased stability of this proteasome activator. SUMOylation-deficient REGγ displayed attenuated ability to degrade p21(Waf//Cip1) due to reduced affinity of the REGγ SUMOylation-defective mutant for p21. Taken together, we report a previously unrecognized mechanism regulating the activity of the proteasome activator REGγ. This regulatory mechanism may enable REGγ to function as a more potent factor in protein degradation with a broader substrate spectrum.