Sentrin-specific protease 3（SENP3）, a member of the desumoylating enzyme family, is known as a redox sensor and could regulate multiple cellular signaling pathways. However, its implication in myocardial ischemia re...Sentrin-specific protease 3（SENP3）, a member of the desumoylating enzyme family, is known as a redox sensor and could regulate multiple cellular signaling pathways. However, its implication in myocardial ischemia reperfusion（MIR） injury is unclear. Here, we observed that SENP3 was expressed and upregulated in the mouse heart depending on reactive oxygen species（ROS） production in response to MIR injury. By utilizing si RNA-mediated cardiac specific gene silencing, SENP3 knockdown was demonstrated to significantly reduce MIR-induced infarct size and improve cardiac function. Mechanistic studies indicated that SENP3 silencing ameliorated myocardial apoptosis mainly via suppression of endoplasmic reticulum（ER） stress and mitochondrial-mediated apoptosis pathways. By contrast, adenovirusmediated cardiac SENP3 overexpression significantly exaggerated MIR injury. Further molecular analysis revealed that SENP3 promoted mitochondrial translocation of dynamin-related protein 1（Drp1） in reperfused myocardium. In addition, mitochondrial division inhibitor-1（Mdivi-1）, a pharmacological inhibitor of Drp1, significantly attenuated the exaggerated mitochondrial abnormality and cardiac injury by SENP3 overexpression after MIR injury. Taken together, we provide the first direct evidence that SENP3 upregulation pivotally contributes to MIR injury in a Drp1-dependent manner, and suggest that SENP3 suppression may hold therapeutic promise for constraining MIR injury.展开更多
白喉毒素突变体蛋白(cross-reacting material 197, CRM197)是一种常用的多糖结合疫苗载体,其在大肠杆菌体系中往往以包涵体形式表达。为了改善通过包涵体复性来制备CRM197而导致的收率低、操作烦琐等不足,研究设计了一种基于SUMO标签...白喉毒素突变体蛋白(cross-reacting material 197, CRM197)是一种常用的多糖结合疫苗载体,其在大肠杆菌体系中往往以包涵体形式表达。为了改善通过包涵体复性来制备CRM197而导致的收率低、操作烦琐等不足,研究设计了一种基于SUMO标签串联的策略,改善重组CRM197的可溶性,同时使用Origami B (DE3)大肠杆菌宿主,成功使重组CRM197蛋白上清液可溶比例提高至95%以上。同时简化了CRM197蛋白的获取途径,经两步Ni亲和层析和一步酶切除去融合标签等手段建立了从破菌上清液中纯化CRM197的方法,获得纯度高于98%的CRM197且收率大于80%。通过圆二色光谱及荧光发射光谱分析等证实纯化所得CRM197空间结构折叠正确。抗原性ELISA实验结果表明所制得的CRM197蛋白与CRM197标准品蛋白抗原性相当。研究结果表明串联双SUMO标签对重组CRM197在大肠杆菌中高效、可溶表达具有促进作用,并建立了快速纯化体系,获得了具有较好免疫原性的CRM197蛋白,为改善重组蛋白在原核表达体系的可溶表达设计提供了新思路。展开更多
蛋白质的翻译后修饰在调节各种细胞过程中的活性和功能中的作用已充分确立。PTM在调节蛋白质构象、定位、活性、稳定性和功能方面起着至关重要作用,并最终诱导了许多基本的生物学功能,包括信号转导、蛋白质转运和定位、蛋白质间相互作...蛋白质的翻译后修饰在调节各种细胞过程中的活性和功能中的作用已充分确立。PTM在调节蛋白质构象、定位、活性、稳定性和功能方面起着至关重要作用,并最终诱导了许多基本的生物学功能,包括信号转导、蛋白质转运和定位、蛋白质间相互作用、细胞分裂和分化,但对于它们之间的相互作用以及它们对同一蛋白靶点的共同调控的研究却很少。本文着眼于一种小泛素化修饰(small ubiquitin related modifier, SUMO),讨论了SUMO化与泛素化、乙酰化以及磷酸化等常见PTM之间的相互联系,并阐述了它们在调节正常生理和发病机制中的作用。展开更多
基金supported by the National Science Fund for Distinguished Young Scholars(No.81625002)the National Natural Science Foundation of China(Nos.81470389,81270282,81330006,81500200,81500221,81070176,81170192,81400261,81600268,and 81601238)+5 种基金the Program of Shanghai Academic Research Leader(18XD1402400)the Shanghai Municipal Education Commission Gaofeng Clinical Medicine Grant Support(No.20152209)Shanghai Shenkang Hospital Development Center(16CR3034A)Shanghai Jiao Tong University(YG2013MS42 and YG2015MS54)Shanghai Jiao Tong University School of Medicine(15ZH1003 and 14XJ10019)the Shanghai Sailing Program(18YF1413000)
文摘Sentrin-specific protease 3（SENP3）, a member of the desumoylating enzyme family, is known as a redox sensor and could regulate multiple cellular signaling pathways. However, its implication in myocardial ischemia reperfusion（MIR） injury is unclear. Here, we observed that SENP3 was expressed and upregulated in the mouse heart depending on reactive oxygen species（ROS） production in response to MIR injury. By utilizing si RNA-mediated cardiac specific gene silencing, SENP3 knockdown was demonstrated to significantly reduce MIR-induced infarct size and improve cardiac function. Mechanistic studies indicated that SENP3 silencing ameliorated myocardial apoptosis mainly via suppression of endoplasmic reticulum（ER） stress and mitochondrial-mediated apoptosis pathways. By contrast, adenovirusmediated cardiac SENP3 overexpression significantly exaggerated MIR injury. Further molecular analysis revealed that SENP3 promoted mitochondrial translocation of dynamin-related protein 1（Drp1） in reperfused myocardium. In addition, mitochondrial division inhibitor-1（Mdivi-1）, a pharmacological inhibitor of Drp1, significantly attenuated the exaggerated mitochondrial abnormality and cardiac injury by SENP3 overexpression after MIR injury. Taken together, we provide the first direct evidence that SENP3 upregulation pivotally contributes to MIR injury in a Drp1-dependent manner, and suggest that SENP3 suppression may hold therapeutic promise for constraining MIR injury.
文摘白喉毒素突变体蛋白(cross-reacting material 197, CRM197)是一种常用的多糖结合疫苗载体,其在大肠杆菌体系中往往以包涵体形式表达。为了改善通过包涵体复性来制备CRM197而导致的收率低、操作烦琐等不足,研究设计了一种基于SUMO标签串联的策略,改善重组CRM197的可溶性,同时使用Origami B (DE3)大肠杆菌宿主,成功使重组CRM197蛋白上清液可溶比例提高至95%以上。同时简化了CRM197蛋白的获取途径,经两步Ni亲和层析和一步酶切除去融合标签等手段建立了从破菌上清液中纯化CRM197的方法,获得纯度高于98%的CRM197且收率大于80%。通过圆二色光谱及荧光发射光谱分析等证实纯化所得CRM197空间结构折叠正确。抗原性ELISA实验结果表明所制得的CRM197蛋白与CRM197标准品蛋白抗原性相当。研究结果表明串联双SUMO标签对重组CRM197在大肠杆菌中高效、可溶表达具有促进作用,并建立了快速纯化体系,获得了具有较好免疫原性的CRM197蛋白,为改善重组蛋白在原核表达体系的可溶表达设计提供了新思路。
文摘蛋白质的翻译后修饰在调节各种细胞过程中的活性和功能中的作用已充分确立。PTM在调节蛋白质构象、定位、活性、稳定性和功能方面起着至关重要作用,并最终诱导了许多基本的生物学功能,包括信号转导、蛋白质转运和定位、蛋白质间相互作用、细胞分裂和分化,但对于它们之间的相互作用以及它们对同一蛋白靶点的共同调控的研究却很少。本文着眼于一种小泛素化修饰(small ubiquitin related modifier, SUMO),讨论了SUMO化与泛素化、乙酰化以及磷酸化等常见PTM之间的相互联系,并阐述了它们在调节正常生理和发病机制中的作用。
文摘炎症小体是一种在炎症反应中发挥重要作用的多蛋白复合体,其激活受到多种因素的调节和影响。小泛素相关修饰物(small ubiquitin-related modifier,SUMO)化是一种翻译后修饰(post-translational modifications,PTMs)的方法,在炎症小体激活的调节过程中也发挥着重要作用。笔者通过对核苷酸结合寡聚化结构域样受体蛋白3(NOD-like receptor family pyrin domain-con-taining protein 3,NLRP3)炎症小体的SUMO化修饰进行系统回顾,阐述多种SUMO修饰分子和SUMO化酶发挥激活NLRP3的生物学作用,总结SUMO化影响NLRP3炎症小体激活的规律,为调节炎症小体激活和治疗炎症小体相关疾病提供新思路。
