SIRT2调控自噬的作用机制研究现状

沉默调节蛋白2(Sirtuin 2,SIRT2)是一种能够调节蛋白质乙酰化修饰的烟酰胺腺嘌呤二核苷酸(NAD+)依赖性去乙酰化酶,是沉默信息调节因子(Sirtuins)家族成员之一,具有多种生物学功能,其在神经退化、细胞分化、葡萄糖和脂质代谢、肿瘤发生、细胞自噬等过程中均发挥调节作用。细胞自噬是一种通过清除异常蛋白或细胞器来维持机体稳态的保护机制,具有促进细胞更新和保持质量的作用。细胞自噬可分为3种主要形式,包括巨自噬、微自噬和分子伴侣介导的自噬,均通过溶酶体来对受损的细胞器、错误折叠和聚集的蛋白质及其他大分子物质进行降解或回收。其中,巨自噬研究最多,分为非选择性巨自噬(即常说的自噬)和选择性巨自噬。线粒体自噬是真核细胞中选择性去除或降解受损和多余线粒体的主要途径,也是SIRT2在细胞中主要调控的一种选择性巨自噬。作者主要综述了SIRT2在自噬及线粒体自噬上的调控作用及机制,以期为后续更深入地研究SIRT2在哺乳动物生理上的更多调控功能及其在各类疾病上的治疗作用提供参考和方向。

SIRT2通过组蛋白H4K8去乳酸化修饰调控巨噬细胞趋化功能

目的·探讨沉默信息调节因子2(silent information regulator 2,SIRT2)通过组蛋白H4第8位赖氨酸(H4K8)去乳酸化修饰对早期感染后巨噬细胞免疫表型的调节作用及相应机制。方法·使用佛波醇-12-肉豆蔻酸酯-13-乙酸酯(phorbol-12-myristate-13-acetate,PMA)诱导人单核细胞白血病THP-1细胞,使其分化为具有巨噬细胞特性的人血单核细胞株(PMA-primed THP-1,pTHP-1),再以脂多糖(lipopolysaccharide,LPS)刺激建立巨噬细胞感染模型。将未经LPS处理的巨噬细胞(pTHP-1)设为对照(CTRL)组,经过LPS处理的巨噬细胞设为感染(LPS)组。通过蛋白质印迹法(Western blotting)检测巨噬细胞中组蛋白乳酸化各位点修饰水平、组蛋白乙酰化各位点修饰水平及SIRT2蛋白水平;通过实时荧光定量PCR(RT-qPCR)方法检测2组间糖酵解限速酶乳酸脱氢酶A(lactate dehydrogenase A,LDHA)、肝脏磷酸果糖激酶(phosphofructokinase liver type,PKFL),糖酵解调剂因子低氧诱导因子1α(hypoxia inducible factor 1α,HIF-1α),以及Sirtuin家族基因和HDAC家族基因表达水平;通过Transwell方法检测巨噬细胞趋化能力;使用慢病毒包装及细胞感染方法建立SIRT2过表达细胞系;使用RNA测序技术(RNA sequencing,RNA-seq)与染色质免疫共沉淀测序技术(chromatin immunoprecipitation sequencing,ChIP-seq)交互分析方法对组蛋白H4第8位赖氨酸乳酸化(lactylation of histone H4 lysine 8,H4K8la)特异性结合的基因进行差异性分析及通路富集分析。结果·相较于CTRL组,LPS组巨噬细胞糖酵解上调,组蛋白H4K8位点乳酸化水平显著增加(P<0.05),而组蛋白其余位点乙酰化水平未见显著变化。所有已知的具有去乳酸化修饰功能的酶中,仅SIRT2在LPS处理后出现显著降低(P<0.05),且SIRT2过表达可显著抑制巨噬细胞中组蛋白H4K8位点的乳酸化水平(P<0.05),但不影响组蛋白H4K8位点的乙酰化水平(P>0.05)。ChIP-seq与RNA-seq交互分析发现,组蛋白H4K8位点乳酸化修饰可调控巨噬细胞趋化相关基因,并且巨噬细胞的趋化能力在SIRT2过表达、H4K8la修饰水平下调后显著下降(P<0.05)。结论·SIRT2可通过去修饰组蛋白H4K8位点乳酸化改变趋化相关靶基因表达,从而降低巨噬细胞趋化能力。靶向SIRT2及H4K8la修饰将有助于控制巨噬细胞介导的炎症反应。

SIRT2在非小细胞肺癌组织中的表达及临床意义

目的探讨沉默信息调节因子2(SIRT2)蛋白在非小细胞肺癌(NSCLC)组织中的表达及与临床病理特征和预后的关系。方法采用免疫组化法检测2011年1月至2013年12月243例NSCLC组织及120例癌旁组织中SIRT2的表达,分析其表达水平与临床病理特征的关系,采用Cox风险比例回归模型分析影响预后的因素。结果SIRT2在NSCLC组织中的高表达率为31.3%(76/243),低于癌旁组织的57.5%(69/120),差异有统计学意义(P<0.001)。SIRT2表达与分化程度和淋巴结转移有关(P<0.05),而与年龄、性别、肿瘤大小和TNM分期均无关(P>0.05)。SIRT2高表达患者的中位无病生存期(DFS)为37.0个月(95%CI:25.3~48.7个月),长于低表达者的31.0个月(95%CI:27.3~34.8个月),差异有统计学意义(P=0.026);SIRT2高表达患者的中位总生存期(OS)为54.0个月(95%CI:40.7~67.3个月),长于低表达者的41.0个月(95%CI:33.6~48.5个月),差异有统计学意义(P=0.023)。Cox多因素回归分析结果显示,分化程度、淋巴结转移是影响NSCLC患者DFS和OS的独立因素(P<0.05),而SIRT2表达对NSCLC患者的预后无独立预测作用(P>0.05)。结论SIRT2在NSCLC组织中低表达,其高表达与NSCLC更好的临床病理特征及预后有关。

Regulation of mitochondrial carrier SLC25A13 on breast cancer cell cycle in vitro

Objective·To investigate the role of mitochondrial solute carrier family 25 member 13(SLC25A13)on breast cancer development.Methods·SLC25A13 mRNA and protein expressions in invasive breast cancer tissues and normal breast tissues were from The Cancer Genome Atlas(TCGA)breast cancer dataset.Survival analysis was conducted online by Kaplan-Meier software.MCF-7 cell line was used for in vitro cell assay.Knockdown of SLC25A13 and sirtuin 2(SIRT2)were conducted by siRNA transfection.Cell viability was measured with trypan blue exclusion.Cell cycle arrest was determined by flow cytometry.The mRNA expression of SLC25A13 and P27 were detected by quantitative PCR.The protein level of SLC25A13,P27 and SIRT2 were detected by Western blotting.Protein half-life of P27 was assessed by Western blotting after cycloheximide treatment.Results·SLC25A13 was up-regulated in invasive breast cancer tissues.High expression of SLC25A13 correlated with poor overall survival and breast cancer recurrence.SLC25A13 knockdown inhibited MCF-7 cell cycle progression.P27 and SIRT2 both accumulated after SLC25A13 knockdown.P27 accumulation resulted from prolonged protein half-life.Knockdown of SIRT2 restored cell cycle arrest as well as P27 accumulation caused by SLC25A13 silencing.Conclusion·High expression of SLC25A13 may promote cell cycle progression via SIRT2 in breast cancer development.

Expression of yeast silencing information regulator 2 in secondary injury of intracerebral hemorrhage

Objective:to investigate the expression of yeast silencing information regulator 2(Sirt2)in the secondary injury of intracerebral hemorrhage(ICH).Methods:twelve Sprague Dawley(SD)rats were randomly divided into a sham group and an ICH group,with six rats in each group.A rat model of ICH was established by injecting collagenase type IV into the right striatum of the rats.The expression of Sirt2 was measured by Western blot and immunohistochemistry after ICH.Result:the behavioral score of the ICH rats was the lowest at 48 h after the operation;therefore the rats at 48 h after surgery were selected as the model rats.The expression of Sirt2 was significantly higher in the striatal tissue of the ICH rats compared with the sham group(P<0.05).Conclusion:the expression of Sirt2 around hematoma in ICH rats decreases,and Sirt2 is expected to become a new target for ICH treatment.

Autophagy in ischemic aged livers

Ischemia/reperfusion(I/R)injury inevitably occurs during liver resection and transplantation.Elderly patients poorly recover from these surgeries.This reduced reparative capacity with aging is causatively associated with decreased mitochondrial function after reperfusion.Mitochondrial autophagy(mitophagy)is a vital cellular process that timely clears abnormal and dysfunctional mitochondria.Impaired or insufficient autophagy can contribute to hepatocyte death after I/R.This review describes our current understanding of I/R injury and highlights new mechanistic correlation between sirtuin 1(SIRT1),mitofusin 2(MFN2),and autophagy in the pathogenesis of age-dependent hypersensitivity to reperfusion injury.The deacetylation of MFN2 by SIRT1 plays a pivotal role in the recovery from reperfusion injury by modulating the onset of autophagy.Targeting the SIRT1-MFN2 axis could be a new therapeutic target to reduce I/R injury in aged livers.