RNA解旋酶和应激颗粒

应激颗粒（stress granules, SGs）是细胞在环境压力刺激下停止蛋白质翻译后,mRNA与多种细胞蛋白组装而成的胞质颗粒结构.RNA 解旋酶家族作为生物体内普遍存在的一类高度保守的蛋白质酶类,参与了RNA代谢各个环节,近年来其家族成员被陆续发现是一类新的SG重要组分.本文综述了RNA解旋酶参与应激颗粒形成过程,RNA解旋酶家族蛋白的结构和其参与应激颗粒形成的研究进展.     

卵磷脂促进小菜蛾颗粒体病毒感染的研究

采用有机溶剂从鸡蛋中提取卵磷脂,与小菜蛾颗粒体病毒混合添食感染2—3龄小菜蛾幼虫,所得数据经统计学方法处理,得到三组死亡率与时间的回归直线方程为y=18.1x-59.8(GV对照)、y=27.23x-99.85(GV+30mg卵磷脂/ml)、y=13.6x-18(GV+70mg卵磷脂/ml)。GV+70mg卵磷脂/ml与GV对照组比较,感染性提高了17％,LT_(50)提前了1.066天。结果表明,卵磷脂对小菜蛾颗粒体病毒感染具有促进作用。     

应激颗粒的形成与生物学意义

真核细胞对外界压力刺激会做出一系列应答反应,如暂停蛋白质翻译系统,从而使细胞能更好地适应环境压力。通过应激颗粒（stress granules,SG）的形成包裹未被翻译的mRNA是该适应性调节的重要方式。研究表明,环境压力导致eIF2α上游激酶的激活从而磷酸化eIF2α,翻译起始受阻,随后,TIA-1、TTP等蛋白迅速与mRNP结合聚集成SG,并在微管蛋白的帮助下进一步向细胞核聚集,形成成熟的SG。当压力消失,SG依赖微管及其动力蛋白进行解聚,释放包裹的mRNA及蛋白。细胞内成熟的SG在转录后调节中发挥重要作用,并且通过其组成蛋白在肿瘤凋亡、病毒侵染、免疫、炎症反应及由蛋白错误折叠引起的疾病中发挥作用。该文首次综述了压力颗粒研究进展,为充分认识SG的病理生理性调节功能提供参考。     

产前束缚应激子代大鼠海马神经颗粒素表达降低

神经颗粒素（neurogranin,NG）是脑特异性突触后蛋白,参与在学习记忆功能中起核心作用的信号转导通路及突触可塑性。本研究旨在探讨产前束缚应激对子代大鼠海马NG表达的影响。连续7d对孕晚期大鼠进行束缚应激,建立产前束缚应激模型,分为对照雌、雄组,应激雌、雄组。采用免疫组化方法观察NG在产前束缚应激子代大鼠海马不同亚区的分布特点;采用蛋白免疫印迹方法检测产前束缚应激子代大鼠海马NG蛋白的表达。结果显示：各组子代大鼠海马各区均有NG蛋白表达,CA1和CA3区表达高于齿状回（dentate gyrus,DG）;应激组雌、雄子代大鼠海马NG的表达明显低于对照组（P〈0．01）,应激组雌性子代比雄性子代减少更显著,对照组雌、雄子代之间无差异。免疫组化与蛋白免疫印迹方法所得结果一致。上述结果表明,NG在产前束缚应激子代大鼠海马表达降低,并且雌性比雄性降低明显,NG对产前束缚应激子代大鼠有差异性调制,NG表达减少可能与产前束缚应激子代大鼠学习记忆能力下降有关。     

应激颗粒在冠状病毒复制中的作用及机制

近年来多种冠状病毒感染后引发患者严重的呼吸道疾病,造成危害人类健康的严重公共卫生事件。新型冠状病毒（SARS-CoV-2）暴发以来,大量研究使得人们对冠状病毒与宿主相互作用机制有更多的了解,其中关于病毒感染后应激颗粒的形成和抗病毒作用也做了大量研究。病毒的RNA和蛋白可以激活宿主细胞内蛋白激酶R(Protein kinase R,PKR)及其下游信号,刺激应激颗粒（Stress granules,SGs）的形成,进而降低病毒在宿主细胞内所需蛋白的翻译水平,抑制病毒复制。然而冠状病毒与宿主长期博弈过程中也衍生出对抗细胞SGs的相应机制,比如利用蛋白与SGs相互作用,来抑制SGs的形成和解聚,逃逸细胞对病毒的抑制作用,保证病毒稳定复制,其中新型冠状病毒就是典型的例子。因此SGs的诱导为抗冠状病毒可能提供一个新型治疗策略。本文对冠状病毒感染抵抗细胞应激颗粒的形成促进其解聚的分子机制进行综述。     

NDV诱导HeLa细胞发生核糖体应激及对翻译起始复合体的影响

[目的]研究新城疫病毒(Newcastle disease virus,NDV) HBUN/LSRC/F3株(以下简称NDV F3)诱导宫颈癌细胞(HeLa)发生核糖体应激后对eIF2α介导的翻译起始复合体eIF4F的调控作用。[方法]流式细胞术及CCK-8检测细胞凋亡;实时荧光定量PCR (quantitative real-time polymerase chain reaction,qRT-PCR)检测c-Myc基因表达;流式细胞术分析细胞周期;Western blotting技术检测c-Myc、RPS7、Bcl-2、NP、eIF4E及eIF2α蛋白的表达;Western blotting和免疫荧光染色技术检测NP、eIF4E蛋白定位。[结果]与阴性对照组相比,NDV F3抑制HeLa细胞增殖并诱导细胞凋亡。细胞周期中G₀/G₁期出现停滞,c-Myc表达呈时间依赖性抑制,c-Myc与Bcl-2蛋白表达量在0-48 h内逐渐下降,NP蛋白在24 h时生成并逐渐增加,RPS7、eIF4E和eIF2α蛋白含量在0-48 h内呈先增加后降低趋势。Western blotting定位分析及激光共聚焦显微镜结果显示NP蛋白主要存在细胞质中,NP与eIF4E存在共定位现象。[结论]NDV F3诱导HeLa细胞凋亡并引发核糖体应激反应,NP与eIF4E相互作用而抑制eIF2α介导的翻译起始复合体eIF4F形成,阻断其与宿主mRNA之间的联系,同时促进NDV F3 mRNA的翻译,最终造成宿主蛋白翻译抑制。     

宿主细胞应答病毒感染的细胞信号转导研究新进展

机体如何识别以及清除入侵的病毒一直是分子免疫学研究的重点.早期的研究揭示,病毒的入侵可诱导表达大量的IFNβ,PKR等抗病毒蛋白分子.这些蛋白质分子通过多种方式造成被侵染细胞表现出特殊的状态或迅速凋亡,从而控制病毒的复制和传播,同时诱导产生大量细胞因子和趋化因子等,启动适应性免疫反应的进程.但是,该领域研究的一个重要瓶颈是对于病毒与宿主细胞相互作用的最早期信号事件了解甚微.近几年的研究工作在先天性免疫系统如何识别早期病毒的入侵方面取得了重大进展.TLR3和RIG-I/MDA5细胞信号转导通路,是最近发现的宿主细胞识别与应答病毒的重要调节机制.它们利用不同的细胞信号转导机制诱导先天性免疫反应,主要参与脊椎动物细胞识别和清除RNA病毒的原发抗感染过程,是机体先天免疫系统的一种重要反应机制,直接影响后续适应性免疫系统的作用.就这些细胞信号转导通路在先天性免疫应答中的研究进展做了概述与展望.     

鼻病毒非结构蛋白2B 诱导细胞发生内质网应激

为探讨鼻病毒非结构蛋白2B诱导内质网应激和细胞凋亡的机制,本研究构建了鼻病毒非结构蛋白2B的真核表达载体p2B‐GFP ,通过转染BHK‐21细胞检测相关标志蛋白的变化情况。结果显示,非结构蛋白2B定位表达于BHK‐21细胞内质网,诱导内质网应激标志蛋白Grp78、CHOP的表达增加,并使活化转录因子6（ATF6）的转录活性增加,还诱导BHK‐21细胞发生核浓缩而凋亡,使凋亡标志蛋白PARP发生降解而减少。结果提示,鼻病毒非结构蛋白2B可诱导细胞发生内质网应激,并经该途径诱导细胞凋亡。     

应激和学习对突触效能的调控机制

长期以来人们知道应激与学习和记忆存在着密切的关系。十多年来的研究进展表明应激调控着长时程增强。然而对应激是否影响到长时程抑制却知之甚少。本文简要的介绍了应有 激,学习对突触可塑性的影响的一些主要实验结果。探讨突中可塑性在神经信息储存中的作用。     

Assessing the genetic component of the susceptibility of mice to viral infections.

Laboratory mice often exhibit wide differences in susceptibility when infected experimentally with viruses. Based on such observations, experiments have been designed to investigate the determinism of these differences at the molecular level, and a few genes that play a major role in the innate mechanisms of defence of the species toward viral aggressions have been characterised. For example, the extraordinary resistance of SJL mice to experimental infections with hepatitis virus strain A59 is the consequence of a structural alteration of a cell adhesion molecule which normally binds to the spikes of the virus, allowing its entry into the cells. If the virus cannot bind to the molecule, or if the molecule is absent, epithelial cells of the intestine and liver are not infected and mice are resistant. In the same way, most--not to say all--laboratory strains of mice are susceptible to infections with orthomyxoviruses or flaviviruses because essential molecules, the synthesis of which is normally triggered by interferon, are defective in these mice. Wild mice, by contrast --probably because they are constantly exposed to natural infections--are resistant. Finally, some mouse strains resist experimental infections by the mouse cytomegalovirus 1 (MCMV-1) because, once infected, these mice synthesise a molecule at the surface of infected cells which allows immediate recognition and killing by natural killer (NK) cells. With the exuberant development of mouse genetics and the constant generation of new mutant alleles, it is likely that many more genes with an impact on the phenotype of resistance or susceptibility will be identified in the forthcoming years. These genes are probably numerous, however, and many of them presumably interact with each other and/or have additive effects. This might slow down progress in our understanding of the innate mechanism of defence.     

Stress granules: RNP-containing cytoplasmic bodies arising in stress: Structure and mechanism of organization

The review considers recent data on stress granules, which are dense RNP-containing cytoplasmic bodies that arise under stress conditions, e.g., in heat shock, UV irradiation, energy depletion, and oxidative stress. There is evidence that stress granules accumulate incomplete initiation complexes containing mRNA associated with proteins, small ribosomal subunits, and some translation initiation factors, and that stress granules are formed when cells are depleted of the ternary complex (eIF2-tRNA^Met-GTP), in particular, upon eIF2A phosphorylation or a decrease in GTP. Large ribosomal subunits and the ternary complex are absent from stress granules. The structural basis of stress granules is known. It is probable, however, that RNA-binding protein TIA-1, which normally occurs in the nucleus, forms prion-like aggregates that serve as scaffolds for other components of stress granules. The cytoskeleton facilitates the accumulation of stress granule components in local cytoplasmic sites. Studies of the formation and composition of stress granules are important for a better understanding of the regulation of translation initiation in vivo and the mechanisms of the cell response to stress factors.     

Stress granules are shock absorbers that prevent excessive innate immune responses to dsRNA

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抗病毒免疫中干扰素与炎症信号通路的交互调控：防御反应与维持稳态

天然免疫是机体通过识别自身或外部危险信号后,为维持体内稳态而逐步建立起来的一系列防御反应,当宿主细胞内的模式识别受体识别胞内病原相关分子模式后激活干扰素(interferon, IFN)、核因子-kappa B (nuclear factor-kappa B, NF-κB)和炎性小体等信号通路。IFNs在天然免疫应答中发挥重要作用,它诱导的抗病毒基因能够通过多种方式抵御病毒的感染,炎症反应则是机体自动的防御反应,能够在病毒感染机体时释放促炎性细胞因子以调控机体的免疫反应,进而发挥抗病毒作用。在病毒感染过程中,IFN信号通路与炎症反应调控网络中的关键分子如NF-κB/RelA、PKR等存在一定的交互作用,此外,IFN信号通路及其产生的细胞因子又影响其他信号通路的活化,进而调控机体的免疫应答以维持自身稳态,它们之间的交互调控失衡将会引起过度炎症反应,导致组织器官的免疫病理损伤,例如SARS-CoV-2感染机体时产生的过度炎症反应。本文综述了机体抗病毒免疫过程中干扰素信号通路与炎症反应之间的交互调控,为研发抗病毒策略提供新思路。     

Bruton’s tyrosine kinase phosphorylates scaffolding and RNA-binding protein G3BP1 to induce stress granule aggregation during host sensing of foreign ribonucleic acids

The Ras-GTPase activating protein SH3 domain-binding protein 1 (G3BP1) plays a critical role in the formation of classical and antiviral stress granules in stressed and virus-infected eukaryotic cells, respectively. While G3BP1 is known to be phosphorylated at serine residues which could affect stress granule assembly, whether G3BP1 is phosphorylated at tyrosine residues and how this posttranslational modification might affect its functions is less clear. Here, we show using immunoprecipitation and immunoblotting studies with 4G10 antibody that G3BP1 is tyrosine-phosphorylated when cells are stimulated with the synthetic double-stranded RNA analog polyinosinic:polycytidylic acid to mimic viral infection. We further demonstrate via co-immunoprecipitation and inhibitor studies that Bruton’s tyrosine kinase (BTK) binds and phosphorylates G3BP1. The nuclear transport factor 2–like domain of G3BP1 was previously shown to be critical for its self-association to form stress granules. Our mass spectrometry, mutational and biochemical cross-linking analyses indicate that the tyrosine-40 residue in this domain is phosphorylated by BTK and critical for G3BP1 oligomerization. Furthermore, as visualized via confocal microscopy, pretreatment of cells with the BTK inhibitor LFM-A13 or genetic deletion of the btk gene or mutation of G3BP1-Y40 residue to alanine or phenylalanine all significantly attenuated the formation of antiviral stress granule aggregates upon polyinosinic:polycytidylic acid treatment. Taken together, our data indicate that BTK phosphorylation of G3BP1 induces G3BP1 oligomerization and facilitates the condensation of ribonucleoprotein complexes into macromolecular aggregates.     

Stress granules counteract senescence by sequestration of PAI‐1

Cellular senescence is a physiological response by which an organism halts the proliferation of potentially harmful and damaged cells. However, the accumulation of senescent cells over time can become deleterious leading to diseases and physiological decline. Our data reveal a novel interplay between senescence and the stress response that affects both the progression of senescence and the behavior of senescent cells. We show that constitutive exposure to stress induces the formation of stress granules (SGs) in proliferative and presenescent cells, but not in fully senescent cells. Stress granule assembly alone is sufficient to decrease the number of senescent cells without affecting the expression of bona fide senescence markers. SG‐mediated inhibition of senescence is associated with the recruitment of the plasminogen activator inhibitor‐1 (PAI‐1), a known promoter of senescence, to these entities. PAI‐1 localization to SGs increases the translocation of cyclin D1 to the nucleus, promotes RB phosphorylation, and maintains a proliferative, non‐senescent state. Together, our data indicate that SGs may be targets of intervention to modulate senescence in order to impair or prevent its deleterious effects.     

Importance of rabies virus nucleoprotein in viral evasion of interferon response in the brain

By using a cultured neuroblastoma cell line, the present authors recently showed that the N protein of virulent rabies virus fixed strain Nishigahara (Ni), but not that of the attenuated derivative Ni‐CE, mediates evasion of induction of type I interferon (IFN). In this study, to determine whether Ni N protein indeed fulfills this function in vivo, the abilities to suppress IFN responses in the mouse brain of Ni‐CE and the virulent chimeric virus CE(NiN), which has the N gene from Ni in the genetic background of Ni‐CE, were compared. It was demonstrated that CE(NiN) propagates and spreads more efficiently than does Ni‐CE in the brain and that IFN response in brains infected with CE(NiN) is weaker than in those infected with Ni‐CE. It was also shown that amino acids at positions 273 and 394 in the N protein, which are known as pathogenic determinants, affect the ability of the viruses to suppress IFN response in the brain. These findings strongly suggest that, in the brain, rabies virus N protein plays important roles in evasion of innate immune responses and thereby in efficient propagation and spread of virus leading to lethal outcomes of infection.     

Viperin triggers ribosome collision-dependent translation inhibition to restrict viral replication

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