子宫球蛋白相关蛋白1的研究进展

子宫球蛋白相关蛋白1(UGRP1)是一种功能尚未完全明确的分泌蛋白,属于子宫球蛋白/Clara细胞分泌蛋白(UG/CCSP)基因超家族,它高度表达于气管,支气管,细支气管,而细支气管中大多数UGRP1阳性的细胞为Clara细胞,UGRP1在氨基酸序列和组织特异性表达上与UG/CCSP相似,且均由肺Clara类似细胞分泌,提示UGRP1与UG/CCSP功能上可能存在一定联系。目前的研究发现,UGRP1可能通过参与炎症反应,在支气管哮喘的发病机制中发挥作用,还可能有与细菌结合发挥清除病原体的作用,具有潜在的免疫活性,它可能通过激活或抑制某些细胞因子的活性而发挥抗炎作用。     

早老素1基因在转染CHO细胞中的表达及其与γ-分泌酶的关系

目的 研究早老素1(PS1)在淀粉样前体蛋白(APP)加工生成β-淀粉样多肽(Ap)过程中的作用及其与γ-分泌酶的关系。方法构建APP和PSI双基因稳定转染的中国仓鼠卵巢(CHO)细胞株,应用免疫沉淀和印迹、脉冲追踪及ELISA方法,检测PS1的表达和代谢半衰期,分析对Aβ分泌的影响及与γ-分泌酶功能的关系。结果PS1转染的CHO细胞(APP-PSI)表达的主要是相对分子质量为45000的全长PS1蛋白,其半衰期短于1h,而其活性片段的N-末端片段和C-末端片段则相对稳定,半衰期接近16h。突变型PS1(M146L)转染细胞分泌的Ap总量与野生型PS1转染细胞没有明显差别,但分泌的Aβ亚型Ap142是未转染PS1或野生型PS1转染细胞分泌的将近2倍。结论 PS1参与了APP加工生成Aβ的过程,突变型PS1(M16L)导致Aβ142的分泌增加,提示PS1可能就是预期的γ-分泌酶。     

SM22α通过调控血管平滑肌细胞PDGF受体再循环参与血管重构过程

目的:本研究拟探讨平滑肌蛋白22α(SM22α)对血管平滑肌细胞(VSMC)表面血小板源性生长因子受体β(PDGFR-β)胞吞的影响,进而揭示SM22α对PDGFR-β活性调控的关键步骤——胞吞和泛素化动态平衡的影响及其对血管重构过程的调控作用。方法:PDGF刺激体外培养大鼠VSMC,考察si RNA敲低SM22α蛋白后不同时点的细胞中,激光共聚焦显微镜观察PDGFR-β在细胞膜分布的异同;利用活细胞工作站实时观察SM22α的表达对PDGFR-β在细胞内内吞体和溶酶体分布的异同;SM22α对c-Cbl或TRAF6等E3泛素连接酶活性的影响;si RNA敲低早期内吞体标志蛋白Rab5、再循环内吞体标志蛋白Rab4和Rab11、多泡体(MVB)标志蛋白Rab7,观察SM22α对PDGFR-β亚细胞定位的变化及其与血管重构的关系。结果:研究发现,SM22α表达下调促进细胞表面的PDGFR-β的再循环过程,使PDGFR-β在细胞表面的分子数显著减少,激活信号分子Akt和p42/44磷酸化,促进PDGF诱导的VSMC生长、增殖和迁移过程,SM22α对PDGFR-β再循环的调控与血管重构过程密切相关。结论:PDGFR-β调控异常诱发的生物学行为改变是心血管疾病的主要细胞和分子生物学基础,我们的结果表明,SM22α可能通过影响PDGFR-β胞吞和泛素化动态平衡而影响其活性的调控,进而参与血管重构的病理生理过程,阐明其分子机制可为发掘基于影响PDGFR-β功能的心血管疾病药物设计提供新靶点。     

整合素相关蛋白与免疫

CD47分子又称整合素相关蛋白,是一种多次跨膜的糖蛋白,在机体的各种细胞及组织上均有表达。CD47分子通过与其整合素配体、信号调节蛋白α(SIRP-α)、凝血酶敏感蛋白(TSP-1)的相互作用,对机体发挥着重要的调控作用。其参与调控细胞的粘附与增殖,激活血小板,抑制红细胞的清除等生理过程,并通过诱导淋巴细胞凋亡,协同刺激T细胞活化,抑制T细胞向Th1型细胞分化,以及影响树突状细胞(DC)的分化成熟及细胞因子的分泌,对免疫系统发挥着重要的调节作用,对诱导免疫耐受、防治移植物抗宿主病(GVHD)及自身免疫性疾病均有重要的意义。     

血管重塑机制研究进展

血管重塑是改善缺血性疾病治疗预后的瓶颈,包括血管壁细胞和细胞外基质结构和形态变化。血流动力学变化通过机械转导机制激活一系列血管生物化学信号,内皮细胞分泌血管活性物质和细胞因子,平滑肌细胞分泌生长因子,可激活多种信号转导途径,通过调控基因表达使血管发生增殖、凋亡、迁移、炎性反应,以及细胞外基质分泌、沉积与降解等变化。本文综述了细胞因子和细胞周期调控蛋白参与血管重塑的研究现状,以及microRNA在调控细胞表型转换、调控炎性反应和调控细胞增殖等方面参与血管重塑。     

整合素相关蛋白与免疫

CD47分子又称整合素相关蛋白，是一种多次跨膜的糖蛋白，在机体的各种细胞及组织上均有表达。CD47分子通过与其整合素配体、信号调节蛋白α(SIRP-α)、凝血酶敏感蛋白(TSP-1)的相互作用，对机体发挥着重要的调控作用。其参与调控细胞的粘附与增殖，激活血小板，抑制红细胞的清除等生理过程，并通过诱导淋巴细胞凋亡，协同刺激T细胞活化，抑制T细胞向Th1型细胞分化，以及影响树突状细胞(DC)的分化成熟及细胞因子的分泌，对免疫系统发挥着重要的调节作用，对诱导免疫耐受、防治移植物抗宿主病(GVHD)及自身免疫性疾病均有重要的意义。     

突触结合蛋白在调节分泌过程中的作用机制

突触结合蛋白(synaptotagm in,Syt)是一类在细胞分泌过程中感受钙离子信号的蛋白质。SytⅠ存在于神经和内分泌细胞的囊泡膜上,被认为是细胞分泌过程中的主要Ca2 感受器,在Ca2 诱导的分泌过程中起关键作用。在哺乳动物细胞中已发现15种亚型,这些Syt蛋白在细胞胞吐和胞吞活动中具有重要的生理功能。Syt的分布、结构以及在细胞蛋白质与膜转运过程中的调节作用及机制的研究具有重要的生理意义。     

调节膜转运的Rab/效应物复合体结构基础

G蛋白是一个种类繁多可与GTP结合而且活性受到GTP调控的超家族。其中的小分子量G蛋白一般只有1个亚单位,而且这些小分子量G蛋白与信息传递,细胞生长与分化、蛋白质合成、合成后的加工及转运机理有密切的关系。Rab蛋白家庭在调控真核细胞膜性细胞器蛋白或其他分子的装运、归类、分泌等活动中起着重要作用。Rab蛋白所调节的功能很广泛,包括囊泡的迁移,及其在膜上特异融合部位的锚定。Rab蛋白在调控突触可塑性(例如神经递质的释放)方面起着决定性的作用。Rab蛋白拥有一个共同的三维折叠结构域,在与GTP结合状态下,它能与下游的多种效应蛋白相结合。如果GTP被水解,这个“开关”区     

神经调节蛋白4调节脂代谢的研究进展

神经调节蛋白4(NRG4)是一种新型脂肪分泌因子,主要在棕色脂肪表达及分泌。NRG4与肝细胞膜上表皮生长因子受体4(ErbB4)特异性结合,激活下游信号通路,抑制脂质的从头合成,增加脂肪酸氧化及生酮作用,减弱肝脂肪变性。NRG4还可调控脂肪组织基因的表达,降低脂肪细胞的胰岛素抵抗,调节脂肪组织中血管重塑,维持脂代谢平衡。NRG4对脂质的有益调节有助于改善与肥胖相关的代谢性紊乱,可能成为非酒精性脂肪肝、糖尿病和动脉粥样硬化等疾病的潜在治疗靶点。     

可诱导共刺激分子诱导骨髓来源不成熟树突状细胞特异性分泌IL-6

目的:分析可诱导共刺激分子(ICOS)可溶性融合蛋白(ICOSIg)能否向不成熟DCs传递逆向信号及其性质。方法:以流式细胞仪结合特异性抗体检测DCs表型分子改变;以ELISA检测培养上清细胞因子变化;以RT-PCR检测各组DCs细胞内细胞因子及受体、趋化因子等mRNA表达水平。结果:ICOSIg或膜锚定ICOS作用于不成熟DCs,均可诱导其高表达MHC-Ⅱ、CD80、CD86和CD83等表型分子;促进DCs特异性分泌IL-6。结论:ICOS作用于不成熟DCs表面的ICOSL可以向DCs细胞传递逆向信号,诱导DCs细胞高分泌IL-6,同时其表面重要的表型分子也上调,可能参与了DCs细胞免疫功能的调节,其信号转导机制可能涉及p38-MAPK通路。     

Drosophila UNC-13 is essential for synaptic transmission.

The UNC-13 protein family has been suggested to be critical for synaptic vesicle dynamics based on its interactions with Syntaxin, Munc-18 and Doc 2alpha. We cloned the Drosophila homolog (Dunc-13) and characterized its function using a combination of electrophysiology and ultrastructural analyses. Dunc-13 contained a C1 lipid-binding motif and two C2 calcium-binding domains, and its expression was restricted to neurons. Elimination of dunc-13 expression abolished synaptic transmission, an effect comparable only to removal of the core complex proteins Syntaxin and Synaptobrevin. Transmitter release remained impaired under elevated calcium influx or application of hyperosmotic saline. Ultrastructurally, mutant terminals accumulated docked vesicles at presynaptic release sites. We conclude that Dunc-13 is essential for a stage of neurotransmission following vesicle docking and before fusion.     

Exocytosis relating proteins in the nervous system

Hypothetical models of the molecular mechanism underlying presynaptic exocytosis were reviewed and the exocytosis relating proteins were categorized into four groups: docking, anchoring, fusion and inhibiting proteins. HPC-1/syntaxin, an axonal membrane protein, was classified as an anchoring protein, not as the vesicle docking protein, because electron microscopic study using cryoimmunogold technique revealed that HPC-1 distributed over the entire axonal membrane, where the synaptic vesicles were not ‘docked’ to the membrane. Since selective toxin or antibody against HPC-1 affected exocytosis, HPC-1 might be a necessary component for the exocytosis, but HPC-1 by itself seemed to have no ability to bind synaptic vesicles to the membrane in vivo. The molecular mechanism for Ca-dependent, rapid exocytosis and possible roles of the exocytosis relating proteins in the neurite morphogenesis are discussed.     

Molecular mechanisms of synaptic vesicle docking and membrane fusion

Studies of the interactions between synaptic proteins are revealing the molecular mechanisms of synaptic vesicle docking and membrane fusion. A 7S complex between the synaptic vesicle proteins synaptotagmin and VAMP, and the plasma membrane proteins syntaxin and SNAP-25 is thought to initiate the docking of vesicles. This is followed by recruitment of the soluble factors α-SNAP and the NSF ATPase to form a ∼20S particle. The hydrolysis of ATP is required for the disassembly of the complex and may ultimately result in the fusion of the membranes. Sequence similarities between syntaxin and neurexins are presented, and a possible role for neurexins in alternative complexes is suggested.     

N type Ca2+ channels and RIM scaffold protein covary at the presynaptic transmitter release face but are components of independent protein complexes

Fast neurotransmitter release at presynaptic terminals occurs at specialized transmitter release sites where docked secretory vesicles are triggered to fuse with the membrane by the influx of Ca²⁺ ions that enter through local N type (CaV2.2) calcium channels. Thus, neurosecretion involves two key processes: the docking of vesicles at the transmitter release site, a process that involves the scaffold protein RIM (Rab3A interacting molecule) and its binding partner Munc-13, and the subsequent gating of vesicle fusion by activation of the Ca²⁺ channels. It is not known, however, whether the vesicle fusion complex with its attached Ca²⁺ channels and the vesicle docking complex are parts of a single multifunctional entity. The Ca²⁺ channel itself and RIM were used as markers for these two elements to address this question. We carried out immunostaining at the giant calyx-type synapse of the chick ciliary ganglion to localize the proteins at a native, undisturbed presynaptic nerve terminal. Quantitative immunostaining (intensity correlation analysis/intensity correlation quotient method) was used to test the relationship between these two proteins at the nerve terminal transmitter release face. The staining intensities for CaV2.2 and RIM covary strongly, consistent with the expectation that they are both components of the transmitter release sites. We then used immunoprecipitation to test if these proteins are also parts of a common molecular complex. However, precipitation of CaV2.2 failed to capture either RIM or Munc-13, a RIM binding partner. These findings indicate that although the vesicle fusion and the vesicle docking mechanisms coexist at the transmitter release face they are not parts of a common stable complex.     

Syntaxin11 serves as a t‐SNARE for the fusion of lytic granules in human cytotoxic T lymphocytes

CTLs kill target cells via fusion of lytic granules (LGs) at the immunological synapse (IS). Soluble N‐ethylmaleimide‐sensitive factor attachment protein receptors (SNAREs) function as executors of exocytosis. The importance of SNAREs in CTL function is evident in the form of familial hemophagocytic lymphohistiocytosis type 4 that is caused by mutations in Syntaxin11 (Stx11), a Qa‐SNARE protein. Here, we investigate the molecular mechanism of Stx11 function in primary human effector CTLs with high temporal and spatial resolution. Downregulation of endogenous Stx11 resulted in a complete inhibition of LG fusion that was paralleled by a reduction in LG dwell time at the IS. Dual color evanescent wave imaging suggested a sequential process, in which first Stx11 is transported to the IS through a subpopulation of recycling endosomes. The resulting Stx11 clusters at the IS then serve as a platform to mediate fusion of arriving LGs. We conclude that Stx11 functions as a t‐SNARE for the final fusion of LG at the IS, explaining the severe phenotype of familial hemophagocytic lymphohistiocytosis type 4 on a molecular level.     

Mechanisms of exocytosis

Catecholamines and peptides secreted from dense-core vesicles (DCVs) of adrenal chromaffin cells regulate a wide variety of physiological processes. For instance, the release of noradrenaline and adrenaline plays a key role in regulating heart rate and blood pressure. Thus understanding the mechanisms of secretory processes of DCVs is crucial for understanding the basis of diseases such as hypertension. DCVs undergo several stages of secretory processing before they are exocytosed. These include docking, priming and triggering of membrane fusion/exocytosis. Molecular studies of DCV exocytosis have identified many proteins critically involved in DCV secretion. These proteins include SNARE proteins, Munc18-1, phosphatidylinositol transfer protein, type I phosphatidylinositol-4-phosphate-5-kinases, NSF, Munc13, CAPS1, synaptotagmins, RalA/RalB GTPases and exocyst proteins. In this article, I will discuss the functions of these proteins within the context of the stages (i.e. docking, priming and triggering of membrane fusion/exocytosis) in DCV secretion.     

The role of actin remodeling in the trafficking of intracellular vesicles, transporters, and channels: focusing on aquaporin-2

Trafficking of the intracellular vesicles and membrane protein incorporated in the vesicles is essential for a variety of basic biological processes. Growing evidence has highlighted the importance of the actin cytoskeleton in the trafficking of synaptic vesicles, secretory granules, transporters, and channels including aquaporin. These trafficking processes require actin remodeling, which is spatiotemporally regulated. Recent researches have come to focus on the motility mechanism of the translocation. In this review, we describe the role of actin at each step of intracellular reservation, exocytosis, docking, fusion with the plasma membrane, and endocytosis, focusing on aquaporin-2 trafficking.     

The kinetic properties of smooth muscle: how a little extra weight makes myosin faster

The contractile properties of smooth muscle (SM) are often described as fast and slow, but the molecular basis for the diversity in contractile properties has yet to be fully elucidated. Studies have shown that the differences in the contractile parameters are seen at the level of the contractile proteins. Experiments have implicated both the splicing of the SM myosin heavy chain (MHC) and the SM myosin essential myosin light chain as possible molecular determinants of the contractile properties of SM. This communication will focus on the role of the 7 aa insert in the smooth muscle MHC in determining the contractile properties of SM and the possible mechanism by which this insert could alter the kinetics of the SM actomyosin ATPase. This revised version was published online in August 2006 with corrections to the Cover Date.     

Mitofusin 2: a mitochondria-shaping protein with signaling roles beyond fusion

Mitochondria are central organelles in metabolism, signal transduction, and programmed cell death. To meet their diverse functional demands, their shape is strictly regulated by a growing family of proteins that impinge on fission and fusion of the organelle. Mitochondrial fusion depends on Mitofusin (Mfn) 1 and 2, two integral outer-membrane proteins. Although MFN1 seems primarily involved in the regulation of the docking and fusion of the organelle, mounting evidence is implicating MFN2 in multiple signaling pathways not restricted to the regulation of mitochondrial shape. Here we review data supporting a role for this mitochondria-shaping protein beyond fusion, in regulating mitochondrial metabolism, apoptosis, shape of other organelles, and even progression through cell cycle. In conclusion, MFN2 appears a multifunctional protein whose biologic function is not restricted to the regulation of mitochondrial shape.     

Smooth Muscle‐Protein Translocation and Tissue Function

Smooth muscle (SM) tissue is a complex organization of multiple cell types and is regulated by numerous signaling molecules (neurotransmitters, hormones, cytokines, etc.). SM contractile function can be regulated via expression and distribution of the contractile and cytoskeletal proteins, and activation of any of the second messenger pathways that regulate them. Spatial‐temporal changes in the contractile, cytoskeletal or regulatory components of SM cells (SMCs) have been proposed to alter SM contractile activity. Ca²⁺ sensitization/desensitization can occur as a result of changes at any of these levels, and specific pathways have been identified at all of these levels. Understanding when and how proteins can translocate within the cytoplasm, or to‐and‐from the plasmalemma and the cytoplasm to alter contractile activity is critical. Numerous studies have reported translocation of proteins associated with the adherens junction and G protein‐coupled receptor activation pathways in isolated SMC systems. Specific examples of translocation of vinculin to and from the adherens junction and protein kinase C (PKC) and 17 kDa PKC‐potentiated inhibitor of myosin light chain phosphatase (CPI‐17) to and from the plasmalemma in isolated SMC systems but not in intact SM tissues are discussed. Using both isolated SMC systems and SM tissues in parallel to pursue these studies will advance our understanding of both the role and mechanism of these pathways as well as their possible significance for Ca²⁺ sensitization in intact SM tissues and organ systems. Anat Rec, 297:1734–1746, 2014. © 2014 Wiley Periodicals, Inc.