胞吐参与植物生物胁迫应答的研究进展

高等植物细胞内囊泡运输介导了膜结构之间的物质运输和信号转导,其中,胞吐调控了运输囊泡向质膜的转运,参与了植物细胞壁形成、细胞分泌和环境响应等多种生物学过程。胞吐可以通过重构及强化细胞壁阻止病原体定殖、分泌抗微生物因子抵御病原体的入侵以及调控植物激素载体蛋白和受体蛋白的极性运输等参与抗病应答。遗传学证据表明胞吐是调控植物生物胁迫响应的重要机制。该文综述胞吐参与植物生物胁迫应答分子机制的研究进展,以期为本领域研究提供参考。     

可兴奋细胞胞吐胞吞的耦联机制

调节型胞吐存在于神经元、内分泌细胞和外分泌细胞等可兴奋细胞以及免疫细胞等特化细胞中,是复杂而精确调节的分泌过程。其作用广泛,与神经信号传递、激素释放、免疫反应等重要的生理活动密切相关。调节型胞吐发生后,在分秒的时间尺度上,可兴奋性细胞快速启动与胞吐发生位置密切相关的调控型胞吞,回收细胞膜脂质和囊泡的膜蛋白,迅速清除分泌位点上由于胞吐而留下的蛋白以利于下一轮的分泌,回收并填充可释放囊泡库,维持细胞膜的平衡。该文先分别介绍可兴奋细胞中胞吐和胞吞的主要模式,然后探讨了它们之间耦联的机制。     

细胞骨架系统调控分泌囊泡与质膜互作的研究进展

胞吐作用是真核生物最基本的细胞活动之一,广泛参与了有机体内的多种生理过程.Exocyst复合体介导的分泌囊泡在质膜的定向栓系以及SNARE(soluble N-ethylmaleimide-sensitive factor attachment protein receptor)蛋白介导的分泌囊泡与质膜的融合过程是胞吐...     

胞泌复合体在植物中的功能研究进展

囊泡运输是真核生物的一种重要的细胞学活动, 广泛参与多种生物学过程。该过程主要包括囊泡形成、转运、拴系及与目的膜融合4个环节。目前已知9种多蛋白亚基拴系复合体参与不同途径的胞内转运过程, 其中, 胞泌复合体(exocyst complex)介导了运输囊泡与质膜的拴系过程。对胞泌复合体调控机制的认识主要源于酵母(Saccharomyces cerevisiae)和动物细胞的研究。近年来, 植物胞泌复合体的研究也取得了较大进展, 初步结果显示复合体在功能方面具有一些植物特异的调控特点, 广泛参与植物生长发育和逆境响应。该文主要综述胞泌复合体在植物中的研究进展, 旨在为植物胞泌复合体功能研究提供参考。     

胞泌复合体在植物中的功能研究进展

囊泡运输是真核生物的一种重要的细胞学活动, 广泛参与多种生物学过程。该过程主要包括囊泡形成、转运、拴系及与目的膜融合4个环节。目前已知9种多蛋白亚基拴系复合体参与不同途径的胞内转运过程, 其中, 胞泌复合体(exocyst complex)介导了运输囊泡与质膜的拴系过程。对胞泌复合体调控机制的认识主要源于酵母(Saccharomyces cerevisiae)和动物细胞的研究。近年来, 植物胞泌复合体的研究也取得了较大进展, 初步结果显示复合体在功能方面具有一些植物特异的调控特点, 广泛参与植物生长发育和逆境响应。该文主要综述胞泌复合体在植物中的研究进展, 旨在为植物胞泌复合体功能研究提供参考。     

钙离子信号对胰岛素分泌的调控

胰岛素的分泌及其分泌的调控是维持机体内葡萄糖平衡的重要机制,胰岛素分泌量的不足会导致非胰岛素依赖的糖尿病的发生.胰岛素包裹在致密核心大囊泡中,胰腺β细胞通过调控致密核心大囊泡的胞吐过程来调节胰岛素的分泌.胞内Ca2 浓度是影响胰岛素分泌的重要因素.胰腺β细胞主要通过质膜上的ATP敏感的钾通道、钙通道和胞内钙库的活动改变胞内Ca2 浓度,从而调控β细胞胰岛素的分泌活动.     

Exocyst复合体的结构与功能研究进展

囊泡运输是真核细胞内细胞器间物质交流的重要手段,主要包括出芽、转运、拴系及膜融合四个环节.拴系因子调控运输囊泡与靶膜的初始接触,建立两者间的连接,并能够促进SNARE介导的膜融合过程.Exocyst是一个保守的八亚基拴系复合体,主要在胞吐过程中介导囊泡与细胞质膜的拴系过程.本文主要介绍exocyst复合体的结构和组装机...     

Ca2+依赖和Ca2+不依赖的囊泡分泌研究进展

Ca2 是促发囊泡胞吐的关键调节因子.最近的研究表明,分泌囊泡和通道之间的空间距离调节囊泡分泌的过程和性质.Ca2 通道开口附近形成的Ca2 微区和Ca2 钠区和囊泡快速递质释放有非常紧密的联系.SNARE蛋白和钙离子传感器synaptotagmins等在触发分泌中起调控作用.同时另有一类不依赖于Ca2 的囊泡分泌存在.Latrotoxin和mastoparan等可以激活这一类不依赖于Ca2 的信号通路,从而触发囊泡释放.本文主要从ca2 对囊泡胞吐的调控作用着手,综述了Ca2 依赖和Ca2 不依赖的囊泡分泌过程和可能的调控机制.     

Ca^2＋依赖和Ca^2＋不依赖的囊泡分泌研究进展

Ca^2＋是促发囊泡胞吐的关键调节因子。最近的研究表明,分泌囊泡和通道之间的空间距离调节囊泡分泌的过程和挂质。Ca^2＋通道开口附近形成的Ca^2＋微区和Ca^2＋钠区和囊泡快速递质释放有非常紧密的联系。SNARE蛋白和钙离子传感器synaptotagmins等在触发分泌中起调控作用。同时另有一类不依赖于Ca^2＋的囊泡分泌存在。Latrotoxin和mastoparan等可以激活这一类不依赖于Ca^2＋的信号通路,从而触发囊泡释放。本文主要从Ca^2＋对囊泡胞吐的调控作用着手,综述了Ca^2＋依赖和Ca^2＋不依赖的囊泡分泌过程和可能的调控机制。     

Synaptotagmin在神经递质释放过程中的作用

神经突触间递质的释放是神经系统完成其生理功能最重要的生物现象之一。在贮存递质的突触囊泡上存在一些神经细胞所特有的囊泡蛋白,如突触素（synapsin）、synaptobrevin和synaptotagmin等。其中synaptotagmins是膜转运蛋白中的一个家族,它们的特征是含有两个钙结合区：C2A和C2B。到目前为止,在哺乳动物中已经发现了15种synaptotagmin同形物（isoforms）。神经递质释放是由Ca^2＋内流以诱导突触囊泡发生胞吐而引起的,Ca^2＋需与细胞内部的Ca^2＋感受器相结合来协同控制囊泡胞吐释放,SynaptotagminⅠ可能作为快速同步释放的Ca^2＋感受器而发挥作用。现在已知synaptotagminⅠ在胞吐和胞吞两个过程中都扮演重要角色。     

Lipid remodelling in neuroendocrine secretion

Neuroendocrine cells secrete hormones and polypeptides through a complex membrane trafficking process that involves the transport of specific organelles, called large dense core secretory granules, from the Golgi apparatus to specialised sites at the plasma membrane where these vesicles are successively exocytosed and recaptured by endocytosis through tightly coupled reactions. The minimal machinery required for exocytosis has been defined as SNARE proteins associated with few accessory proteins. On the other side, clathrin and dynamin constitute major components of some of the most important endocytotic pathways. Although many protein contributors of both exocytosis and endocytosis are now identified, their actual interplay is not well resolved. Furthermore, the necessary tight coupling of exocytosis and compensatory endocytosis to maintain membrane homeostasis in neuroendocrine cells is far from being understood. In this review, we focus on the more recently identified role of lipids in these important processes that are above all membrane remodelling events.     

Actin remodeling to facilitate membrane fusion

Actin and its associated proteins participate in several intracellular trafficking mechanisms. This review assesses recent work that shows how actin participates in the terminal trafficking event of membrane bilayer fusion. A recent flurry of reports defines a role for Rho proteins in membrane fusion and also demonstrates that this role is distinct from any vesicle transport mechanism. Rho proteins are well known to govern actin remodeling, which implicates this process as a condition of membrane fusion. A small but significant body of work examines actin-regulated events of intracellular membrane fusion, exocytosis and endocytosis. In general, actin has been shown to act as a negative regulator of exocytosis. Cortical actin filaments act as a barrier that requires transient removal to allow vesicles to undergo docking at the plasma membrane. However, once docked, F-actin synthesis may act as a positive regulator to give the final stimulus to drive membrane fusion. F-actin synthesis is clearly needed for endocytosis and intracellular membrane fusion events. What may seem like dissimilar results are perhaps snapshots of a single mechanism of membranous actin remodeling (i.e. dynamic disassembly and reassembly) that is universally needed for all membrane fusion events.     

Sweeping model of dynamin activity. Visualization of coupling between exocytosis and endocytosis under an evanescent wave microscope with green fluorescent proteins

Vesicle recycling through exocytosis and endocytosis is mediated by a coordinated cascade of protein-protein interactions. Previously, exocytosis and endocytosis were studied separately so that the coupling between them was understood only indirectly. We focused on the coupling of these processes by observing the secretory vesicle marker synaptobrevin and the endocytotic vesicle marker dynamin I tagged with green and red fluorescent proteins under an evanescent wave microscope in pheochromocytoma cells. In control cells, many synaptobrevin-expressing vesicles were found as fluorescent spots near the plasma membrane. Upon electrical stimulation, many of these vesicles showed an exocytotic response as a transient increase in fluorescence intensity followed by their disappearance. In contrast, fluorescent dynamin appeared as clusters increasing slowly in number upon stimulation. The clusters of fluorescent dynamin moved around beneath the plasma membrane for a significant distance. Simultaneous observations of green fluorescent dynamin and red fluorescent synaptobrevin indicated that more than 70% of the exocytotic responses of synaptobrevin had no immediate dynamin counterpart at the same site. From these findings it was concluded that dynamin-mediated recycling is not directly coupled to exocytosis but rather completed by a scanning movement of dynamin for the sites of invaginating membrane destined to endocytosis.     

Endocytosis in plants: Peculiarities and roles in the regulated trafficking of plant metal transporters

The removal of transmembrane proteins from the plasma membrane via endocytosis has emerged as powerful strategy in the regulation of receptor signalling and molecule transport. In the last decade, IRON‐REGULATED TRANSPORTER1 (IRT1) has been established as one of the key plant model proteins for studying endomembrane trafficking. The use of IRT1 and additional other metal transporters has uncovered novel factors involved in plant endocytosis and facilitated a better understanding of the role of endocytosis in the fine balancing of plant metal homoeostasis. In this review, we outline the specifics of plant endocytosis compared to what is known in yeast and mammals, and based on several examples, we demonstrate how studying metal transport has contributed to extending our knowledge of endocytic trafficking by shedding light on novel regulatory mechanisms and factors.     

SNAP25, but not syntaxin 1A, recycles via an ARF6-regulated pathway in neuroendocrine cells

Soluble N-ethylmaleimide-sensitive fusion protein attachment protein receptor (SNARE) proteins mediate cellular membrane fusion events and provide a level of specificity to donor-acceptor membrane interactions. However, the trafficking pathways by which individual SNARE proteins are targeted to specific membrane compartments are not well understood. In neuroendocrine cells, synaptosome-associated protein of 25 kDa (SNAP25) is localized to the plasma membrane where it functions in regulated secretory vesicle exocytosis, but it is also found on intracellular membranes. We identified a dynamic recycling pathway for SNAP25 in PC12 cells through which plasma membrane SNAP25 recycles in approximately 3 h. Approximately 20% of the SNAP25 resides in a perinuclear recycling endosome-trans-Golgi network (TGN) compartment from which it recycles back to the plasma membrane. SNAP25 internalization occurs by constitutive, dynamin-independent endocytosis that is distinct from the dynamin-dependent endocytosis that retrieves secretory vesicle constituents after exocytosis. Endocytosis of SNAP25 is regulated by ADP-ribosylation factor (ARF)6 (through phosphatidylinositol bisphosphate synthesis) and is dependent upon F-actin. SNAP25 endosomes, which exclude the plasma membrane SNARE syntaxin 1A, merge with those derived from clathrin-dependent endocytosis containing endosomal syntaxin 13. Our results characterize a robust ARF6-dependent internalization mechanism that maintains an intracellular pool of SNAP25, which is compatible with possible intracellular roles for SNAP25 in neuroendocrine cells.     

Vesicle trafficking dynamics and visualization of zones of exocytosis and endocytosis in tobacco pollen tubes

Pollen tubes are one of the fastest growing eukaryotic cells.Rapid anisotropic growth is supported by highly active exocytosisand endocytosis at the plasma membrane, but the subcellularlocalization of these sites is unknown. To understand molecularprocesses involved in pollen tube growth, it is crucial to identifythe sites of vesicle localization and trafficking. This reportpresents novel strategies to identify exocytic and endocyticvesicles and to visualize vesicle trafficking dynamics, usingpulse-chase labelling with styryl FM dyes and refraction-freehigh-resolution time-lapse differential interference contrastmicroscopy. These experiments reveal that the apex is the siteof endocytosis and membrane retrieval, while exocytosis occursin the zone adjacent to the apical dome. Larger vesicles areinternalized along the distal pollen tube. Discretely sizedvesicles that differentially incorporate FM dyes accumulatein the apical, subapical, and distal regions. Previous workestablished that pollen tube growth is strongly correlated withhydrodynamic flux and cell volume status. In this report, itis shown that hydrodynamic flux can selectively increase exocytosisor endocytosis. Hypotonic treatment and cell swelling stimulatedexocytosis and attenuated endocytosis, while hypertonic treatmentand cell shrinking stimulated endocytosis and inhibited exocytosis.Manipulation of pollen tube apical volume and membrane remodellingenabled fine-mapping of plasma membrane dynamics and definedthe boundary of the growth zone, which results from the orchestratedaction of endocytosis at the apex and along the distal tubeand exocytosis in the subapical region. This report providescrucial spatial and temporal details of vesicle traffickingand anisotropic growth. Key words: Endocytosis; exocytosis, hydrodynamics, lipophilic FM dyes, pollen tube growth, vesicle trafficking Received 14 September 2007; Revised 23 November 2007 Accepted 7 January 2008     

Trafficking of the plasma membrane gamma-aminobutyric acid transporter GAT1. Size and rates of an acutely recycling pool

Plasma membrane neurotransmitter transporters rapidly traffic to and from the cell surface in neurons. This trafficking may be important in regulating neuronal signaling. Such regulation will be subject to the number of trafficking transporters and their trafficking rates. In the present study, we define an acutely recycling pool of endogenous gamma-aminobutyric acid transporters (GAT1) in cortical neurons that comprises approximately one-third of total cellular GAT1. Kinetic analysis of this pool estimates exocytosis and endocytosis time constants of 1.6 and 0.9 min, respectively, and thus approximately one-third of the recycling pool is plasma membrane resident in the basal state. Recent evidence shows that GAT1 substrates, second messengers, and interacting proteins regulate GAT1 trafficking. These triggers could act by altering trafficking rates or by changing the recycling pool size. In the present study we examine three GAT1 modulators. Calcium depletion decreases GAT1 surface expression by diminishing the recycling pool size. Sucrose increases GAT1 surface expression by blocking clathrin- and dynamin-dependent endocytosis, but it does not change the recycling pool size. Protein kinase C decreases surface GAT1 expression by increasing the endocytosis rate, but it does not change the exocytosis rate or the recycling pool size. Based upon estimates of GAT1 molecules in cortical boutons, the present data suggest that approximately 1000 transporters comprise the acutely recycling pool, of which 300 are on the surface in the basal state, and five transporters insert into the plasma membrane every second. This insertion could represent the fusion of one transporter-containing vesicle.     

参与植物膜泡运输的蛋白质家族及功能研究进展

真核细胞的内吞和分泌途径中蛋白质和脂类的运输主要由膜泡运输介导。参与膜泡运输的蛋白质家族包括SNARE蛋白家族、RAB蛋白家族、被膜蛋白复合体、Sec1蛋白家族、Arf蛋白家族。这些蛋白质家族在进化中高度保守,并且在植物中已经鉴定了许多哺乳动物和酵母蛋白的同源物。近年来一些研究发现这些蛋白质不仅仅调节植物细胞的膜泡运输,还影响植物的许多生理活动和功能,例如向重性生长、胞质分裂、激素极性运输、气孔运动以及抗病性等。现主要阐述迄今在植物中研究这五类蛋白质家族功能的最新进展。     

Endosomal functions in plants

Plant endosomes are highly dynamic organelles that are involved in the constitutive recycling of plasma membrane cargo and the trafficking of polarized plasma membrane proteins such as auxin carriers. In addition, recent studies have shown that surface receptors such as the plant defense-related FLS2 receptor and the brassinosteroid receptor BRI1 appear to signal from endosomes upon ligand binding and internalization. In yeast and mammals, endosomes are also known to recycle vacuolar cargo receptors back to the trans Golgi network and sort membrane proteins for degradation in the vacuole/lysosome. Some of these sorting mechanisms are mediated by the retromer and endosomal sorting complex required for transport (ESCRT) complexes. Plants contain orthologs of all major retromer and ESCRT complex subunits, but they have also evolved variations in endosomal functions connected to plant-specific features such as the diversity of vacuolar transport pathways. This review focuses on recent studies in plants dealing with the regulation of endosomal recycling functions, architecture and formation of multivesicular bodies, ligand-mediated endocytosis and receptor signaling from endosomes as well as novel endosomal markers and the function of endosomes in the transport and processing of soluble vacuolar proteins.     

Vesicle-associated membrane protein 2 mediates trafficking of α5β1 integrin to the plasma membrane

Integrins are major receptors for cell adhesion to the extracellular matrix (ECM). As transmembrane proteins, the levels of integrins at the plasma membrane or the cell surface are ultimately determined by the balance between two vesicle trafficking events: endocytosis of integrins at the plasma membrane and exocytosis of the vesicles that transport integrins. Here, we report that vesicle-associated membrane protein 2 (VAMP2), a SNARE protein that mediates vesicle fusion with the plasma membrane, is involved in the trafficking of α5β1 integrin. VAMP2 was present on vesicles containing endocytosed β1 integrin. Small interfering RNA (siRNA) silencing of VAMP2 markedly reduced cell surface α5β1 and inhibited cell adhesion and chemotactic migration to fibronectin, the ECM ligand of α5β1, without altering cell surface expression of α2β1 integrin or α3β1 integrin. By contrast, silencing of VAMP8, another SNARE protein, had no effect on cell surface expression of the integrins or cell adhesion to fibronectin. In addition, VAMP2-mediated trafficking is involved in cell adhesion to collagen but not to laminin. Consistent with disruption of integrin functions in cell proliferation and survival, VAMP2 silencing diminished proliferation and triggered apoptosis. Collectively, these data indicate that VAMP2 mediates the trafficking of α5β1 integrin to the plasma membrane and VAMP2-dependent integrin trafficking is critical in cell adhesion, migration and survival.