SUMO化修饰在病毒感染中的作用研究进展

Small ubiquitin-related modifier（SUMO）是一类重要的类泛素蛋白,其修饰过程与泛素类似,但功能不尽相同,SUMO通过对底物蛋白的修饰来调节蛋白间的相互作用、定位及稳定性等功能。近年来,包括一些病毒蛋白在内的大量底物蛋白陆续被发现,SUMO化作为一种重要的蛋白质翻译后修饰,在病毒对宿主细胞的感染中既可发挥促进病毒感染的作用,也可发挥抑病毒感染的作用。     

蛋白质泛素化修饰及其在脓毒症中的研究进展

蛋白质在机体生命活动中扮演关键角色,其降解途径中,泛素-蛋白酶体途径(UPS)通过泛素化修饰调控蛋白降解,同时通过共价修饰底物蛋白参与细胞生理活动。去泛素化酶(DUBs)对蛋白质泛素化具有平衡作用,通过移除泛素化修饰来维持泛素化修饰的动态平衡,它们在脓毒症中发挥着关键作用,通过调控炎症因子的表达,影响机体的炎症反应。脓毒症是导致患者重症监护的主要疾病,涉及复杂炎症反应。蛋白质泛素化修饰参与脓毒症信号转导,影响核因子-κB(NF-κB)信号通路和NOD样受体蛋白3炎症小体(NLRP3)活化。这些调控机制影响细胞内炎症因子的释放,进而影响机体炎症反应的强度和时机。本文探讨了近年来与脓毒症相关关键蛋白的泛素化及去泛素化机制,以期为脓毒症的治疗提供新的靶点和策略。     

tau蛋白异常翻译后修饰在阿尔茨海默病中的作用

在阿尔茨海默病(Alzheimer's disease,AD)中微管相关蛋白tau能够产生许多异常翻译后修饰并聚集形成配对螺旋丝(paired helical filament,PHF).这些tau的修饰包括过磷酸化、异常糖基化、截断等,其中,过磷酸化和异常糖基化是阿尔茨海默氏病等神经退行性疾病神经元纤维化的主要分子发病机制.     

蛋白质磷酸化修饰在多聚谷氨酰胺疾病中的研究进展

多聚谷氨酰胺(polyglutamine,polyQ)疾病是一大组常见的神经退行性疾病,疾病的发生源于致病基因编码区CAG三核苷酸重复扩展突变导致基因的编码蛋白--polyQ蛋白产生多聚谷氨酰胺扩展突变.polyQ疾病的发病机制目前虽然尚未得到完全阐明,但越来越多的研究表明蛋白质的磷酸化修饰在亨廷顿舞蹈病、齿状核红核苍白球路易氏体萎缩症、延髓脊肌萎缩症、遗传性脊髓小脑型共济失调1型、遗传性脊髓小脑型共济失调3型/马查多.约瑟夫病等疾病的发生发展中发挥了重要的作用.     

炎症反应中的蛋白质翻译后修饰

炎症相关的信号转导蛋白、转录因子、炎性介质、组蛋白等可在炎症发生发展过程中发生磷酸化、乙酰化、泛素化、苏素化、甲基化等一系列翻译后修饰.这些化学修饰可高效调节相关蛋白质的功能活性及基因表达水平,不同化学修饰之间还可相互作用,共同影响炎症的发生、发展与转归;而异常的翻译后修饰与炎症相关性疾病关系密切.     

不同疱疹病毒相关蛋白与SUMO修饰系统的相互作用

小泛素化相关修饰物(small ubiquitin-related modifier,SUMO)修饰是一种与泛素化修饰具有相似酶促反应过程的翻译后修饰.近年来,研究者相继发现许多病毒蛋白能发生SUMO修饰,特别是疱疹病毒相关蛋白.研究表明,SUMO修饰能影响一些病毒蛋白的转录调控活性.此外,一些定位于核内功能性结构PODs(PML oncogenic domains)的病毒蛋白还能影响SUMO-1对PODs主要成分PML及Sp100的修饰并导致PODs解聚.因此了解SUMO修饰如何改变病毒与细胞相互作用的生物学特件对研究病毒的复制增殖机制具有重要的意义.本文就疱疹病毒相关蛋白与SUMO修饰系统相互作用的分子机制及功能影响进行综述.     

蛋白质SUMO化和泛素化修饰的关系

泛素化和SUMO化是蛋白质翻译后修饰的重要方式,广泛参与调节蛋白质功能和细胞生命活动各个环节.多聚泛素化降解蛋白质,而SUMO化主要调节蛋白质的相互作用和定位等.在不同情况下,SUMO化和泛素化既可协同调节蛋白质功能,也可相互拮抗. 最近研究发现,某些底物的SUMO化能够激活体内一类新发现的SUMO依赖的泛素连接酶,启动泛素-蛋白酶体途径降解底物, 导致蛋白质SUMO化和泛素化的关系进一步精细化和复杂化.     

P53翻译后修饰研究进展

肿瘤抑制基因p53在肿瘤发生中发挥着重要作用。P53蛋白的翻译后修饰及其与多种细胞蛋白间相互作用使P53蛋白呈现功能多样性。P53的翻译后修饰不是单个位点的修饰而是包括磷酸化、乙酰化、泛素化及SUMO化作用的多位点修饰。翻译后修饰对P53功能至关重要,更可能与某些肿瘤的发生密切相关。发生在蛋白质水平上的P53的功能性灭活是没有发生p53基因突变肿瘤发生的重要机制之一。     

Tau pathology modulates Pin1 post-translational modifications and may be relevant as biomarker

A prerequisite to dephosphorylation at Ser–Pro or Thr–Pro motifs is the isomerization of the imidic peptide bond preceding the proline. The peptidyl-prolyl cis/trans isomerase named Pin1 catalyzes this mechanism. Through isomerization, Pin1 regulates the function of a growing number of targets including the microtubule-associated tau protein and is supposed to be deregulated Alzheimer's disease (AD). Using proteomics, we showed that Pin1 is posttranslationally modified on more than 5 residues, comprising phosphorylation, N-acetylation, and oxidation. Although Pin1 expression remained constant, Pin1 posttranslational two-dimensional pattern was modified by tau overexpression in a tau-inducible neuroblastoma cell line, in our THY-Tau22 mouse model of tauopathy as well as in AD. Interestingly, in all of these systems, Pin1 modifications were very similar. In AD brain tissue when compared with control, Pin1 is hyperphosphorylated at serine 16 and found in the most insoluble hyperphosphorylated tau fraction of AD brain tissue. Furthermore, in all tau pathology conditions, acetylation of Pin1 may also contribute to the differences observed. In conclusion, Pin1 displays several posttranslational modifications, which are specific in tauopathies and may be useful as biomarker.     

Promyelocytic leukaemia (PML) protein expression in human placenta and choriocarcinoma

Promyelocytic leukaemia (PML) protein, the product of the pml gene, is heterogeneously expressed in various normal and neoplastic tissues, and the fusion of the pml gene with retinoic acid receptor-alpha is believed to be a central mechanism in acute PML tumourigenesis. As PML is important for controlling major cellular processes, such as growth and differentiation, it is believed that it plays an important role during human gestation. The human placenta is a critical organ for the maintenance of gestation, but the expression pattern and functional significance of PML in the placenta have not been documented. The present study has therefore investigated the expression of PML in the human placenta and in choriocarcinoma, and has observed the biological effects following the overexpression of PML in choriocarcinoma cell lines (BeWo and JEG-3). In the human placenta, PML expression was readily found in villous stromal fibroblasts, capillary endothelial cells, Hofbauer cells, and occasionally in amnion cells. Moreover, immunoblotting of placental lysates demonstrated increased PML expression with increasing gestation. Interestingly, PML expression was confined to intermediate trophoblasts and syncytiotrophoblastic giant cells at the placental site (placental site giant cells) in the trophoblastic cell population. Intermediate trophoblasts at non-placental sites, and villous cytotrophoblasts and syncytiotrophoblasts consistently did not express PML. Further screening of PML expression in hydatidiform moles (n = 4) and choriocarcinomas (n = 7) also revealed selective PML expression in intermediate trophoblastic cells and syncytiotrophoblastic cells, but not in the cytotrophoblastic populations, which corresponds well with observations in the placental bed. Adenoviral transduction of PML resulted in a marked reduction in cell growth in both choriocarcinoma cell lines, which was associated with increased apoptosis. The findings of the present study strongly suggest that PML plays an important role in human placental development and growth, and in the pathobiology of trophoblasts and trophoblastic neoplasia.     

重型再生障碍性贫血患者血清诱导淋巴细胞凋亡及PML蛋白表达

用免疫荧光染色及Western blot检测重型再障患者血清对正常人淋巴细胞凋亡的促进作用及早幼粒细胞白血病(PML)蛋白表达的诱导作用,并进一步观察Caspase3,8抑制剂作用前后这两者的变化,以探讨再生障碍性贫血(再障)患者淋巴细胞凋亡增加的机制.结果显示,10例重型再障患者血清作用后,淋巴细胞的凋亡率较正常人血清作用后明显增加(P＜0.05).同时,PML蛋白也明显高表达(P＜0.001),两者呈正相关(r=0.919,P＜0.001).caspase8抑制剂可部分阻断再障患者血清的这种作用,但caspase3抑制剂不能阻断这种作用.以上结果提示重型再障患者血清可诱导淋巴细胞PML蛋白的高表达,从而导致淋巴细胞凋亡,caspase8可能参与了这一过程.     

Nuclear body formation and PML body remodeling by the human cytomegalovirus protein UL35

The human cytomegalovirus (HCMV) UL35 gene encodes two proteins, UL35 and UL35a. Expression of UL35 in transfected cells results in the formation of UL35 nuclear bodies that associate with promyelocytic leukemia (PML) protein. PML forms the basis for PML nuclear bodies that are important for suppressing viral lytic gene expression. Given the important relationship between PML and viral infection, we have further investigated the association of UL35 with PML bodies. We demonstrate that UL35 bodies form independently of PML and subsequently recruit PML, Sp100 and Daxx. In contrast, UL35a did not form bodies; however, it could bind UL35 and inhibit the formation of UL35 bodies. The HCMV tegument protein pp71 promoted the formation of UL35 bodies and the cytoplasmic localization of UL35a. Similarly, UL35a shifted pp71 to the cytoplasm. These results indicate that the interplay between UL35, UL35a and pp71 affects their subcellular localization and likely their functions throughout infection.     

腺病毒介导的PML(NLS-)过表达对白血病NB4细胞增殖和凋亡的影响

目的 利用腺病毒介导PML(NLS-)的过表达,探讨PML(NLS-)对NB4白血病细胞增殖凋亡的影响.方法 重组腺病毒质粒Ad-PML(NLS-)经Pac Ⅰ酶切线性化后转染AD293细胞,获得重组腺病毒Ad-PML(NLS-),经4轮扩增后,测定重组腺病毒滴度;重组腺病毒感染NB4细胞,荧光显微成像和流式细胞术检测感染效率,RT-PCR法和Western blot法检测NB4细胞中PML(NLS-)的转录及表达水平,MTT实验观察细胞增殖,流式细胞术分析细胞周期及凋亡.结果 重组腺病毒质粒Ad-PML(NLS-)经酶切和测序鉴定正确,经4轮扩增病毒滴度为1 ×1010pfu/mL;重组腺病毒Ad-PML(NLS-)感染NB4细胞效率达75％,PML(NLS-)基因可在NB4细胞中高效表达;NB4细胞增殖活力明显高于未感染组和空病毒载体感染组(P＜0.05),S期细胞比例明显增高(P＜0.05),G2期细胞比例明显减少(P＜0.05);实验组细胞凋亡率明显低于空病毒感染组(P＜0.05).结论 PML(NLS-)过表达能够促进白血病NB4细胞的体外增殖能力,抑制其凋亡.     

The functional heterogeneity of eosinophil cationic protein is determined by a gene polymorphism and post-translational modifications

BACKGROUND: The eosinophil is a cytotoxic cell and takes part in parasite killing and tissue-destructive processes by secretion of proteins such as eosinophil cationic protein (ECP). A polymorphism was demonstrated in the ECP gene, giving rise to a substitution of arginine at position 97 with threonine. This polymorphism is related to disease development. OBJECTIVE: To investigate the functional and molecular heterogeneity of native ECP and the functional consequences of the replacement of arginine with a threonine. METHODS: ECP was purified from healthy blood donors by gel filtration, ion-exchange chromatography and reversed-phase chromatography. Recombinant ECPs i.e. rECP 97(arg) and rECP 97(thr) were produced by the pFASTBAC baculovirus expression system. The cytotoxic activity was determined against an erythroleukaemia or a small cell lung cancer cell line. RESULTS: Native ECP was purified to apparent homogeneity and showed a considerable molecular heterogeneity and a corresponding functional heterogeneity with respect to cytotoxic activity. After reduction, the native cytotoxic ECP showed three bands on sodium dodecylsulphate polyacrylamide gel electrophoresis : one major band at 18-20 kDa and two minor bands at about 10 and 5 kDa, respectively. The 5 kDa contained two masses differing with 56.2 Da, which corresponds to the difference in molecular masses of arginine and threonine. rECP 97(arg) was cytotoxic in contrast to rECP97(thr). Deglycosylation with N-glycosidase F did not affect the cytotoxic activity of native ECP to any measurable extent nor the activity of rECP 97(arg), whereas rECP 97(thr) achieved cytotoxic activity. The RNase activities of the recombinant and native ECPs were similar. CONCLUSION: We conclude that ECP is present in several molecular forms with varying biological activities. Some of this functional heterogeneity is based on the genetic polymorphism of the ECP gene and some on post-translational modifications. In subjects carrying the ECP 97(thr) variant, the cytotoxic activity may be disguised by N-linked glycosylation of the active site.     

The role of WRN in DNA repair is affected by post-translational modifications

Werner syndrome (WS) is an autosomal recessive progeroid disease characterized by genomic instability. WRN gene encodes one of the RecQ helicase family proteins, WRN, which has ATPase, helicase, exonuclease and single stranded DNA annealing activities. There is accumulating evidence suggesting that WRN contributes to the maintenance of genomic integrity through its involvement in DNA repair, replication and recombination. The role of WRN in these pathways can be modulated by its post-translational modifications in response to DNA damage. Here, we review the functional consequences of post-translational modifications on WRN as well as specific DNA repair pathways where WRN is involved and discuss how these modifications affect DNA repair pathways.