Regulation of membrane fusion by a novel Sec1/Munc18-associated protein

新型 Sec1/Munc18 相关蛋白对膜融合的调节

• 批准号: BB/H024867/1
• 负责人: Michael Blatt
• 金额: $ 60.63万
• 依托单位: University of Glasgow
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2011
• 资助国家: 英国
• 起止时间: 2011 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=BB%2FH024867%2F1
• 关键词: Regulation; membrane; fusion; novel; Sec1

SNARE proteins are central components of a well-defined mechanism for the delivery of vesicles carrying membrane and soluble cargo between compartments within eukaryotic cells. Vesicle traffic contributes to neurotransmitter release in nerves, to cell wall delivery and budding in yeast, and is essential for cellular homeostasis, growth and development in plants. Cognate SNARE proteins localise to vesicle and target membranes, and assembly of functional SNARE complexes is sufficient to drive membrane fusion. SNAREs also bind other protein partners, specialised to align vesicle fusion within certain physiological roles. SNARE interactions with ion channels in animals appear to facilitate electrical signalling and neuroendocrine secretion, and we have shown that interaction between the Arabidopsis plasma membrane SNARE SYP121 and the ion channel subunit KC1 affects transport of the osmotically-active K+ ion. An obvious potential corollary of this is that binding between SNAREs and ion channels is important in regulating vesicle fusion. Despite the fundamental nature of the process, we have little understanding of the molecular mechanisms that couple cellular volume with osmotic solute transport in plants or other walled eukaryotic cells. Indeed, how walled cells regulate transport of osmotically-active solutes (especially of K+) in parallel with cell volume - whether reversible as in guard cells or during irreversible expansive growth - remains a matter of considerable debate. This proposal builds on significant recent findings we have made following on our identification of the SYP121-KC1 protein complex and its role in K+ channel regulation: (1) we have identified the site on the SNARE protein SYP121 that binds the channel subunit KC1, and (2) we have demonstrated that KC1 competes with the Sec1/Munc18 (SM) protein KEULE for binding to SYP121. SM proteins are key regulators of membrane vesicle fusion in all eukaryotes, but their precise role in controlling SNARE-mediated membrane fusion remains unclear and a topic of intense research. Our findings point to a link between SM protein function and a SNARE-channel interaction. Not only do they offer the first evidence of a mechanism that directly couples osmotic solute transport with vesicle traffic to control cell expansion in walled eukaryotes, but they also support a new model for SM-regulated membrane traffic. Our working hypothesis is that the SNARE-K+ channel interaction of SYP121 and KC1 serves as a 'molecular governor', analogous to the mechanical invention James Watt employed in moderating the turnover rate of his steam engines, to coordinate vesicle traffic (and cell expansion) with uptake of the osmotically-active K+ ion. We now propose to test various aspects of this hypothesis. We aim to fully characterize the binding of SYP121 to KC1 and KEULE in order to build up a molecular map of the interactions. We also propose to examine the consequences of selectively disrupting the interactions between SYP121, KC1 and KEULE on SNARE complex assembly, membrane traffic and channel-mediated K+ transport. Our multidisciplinary approach will not only further our understanding of the link between osmotic solute transport and control of cell volume, but is also likely to provide a novel paradigm for linking membrane traffic with other physiological processes.

SNARE蛋白是真核细胞内在区室之间递送携带膜和可溶性货物的囊泡的明确机制的核心组分。囊泡运输有助于神经中的神经递质释放，酵母中的细胞壁递送和出芽，并且对于植物中的细胞内稳态、生长和发育至关重要。同源SNARE蛋白定位于囊泡和靶膜，功能性SNARE复合物的组装足以驱动膜融合。SNARE还结合其他蛋白质伴侣，专门在某些生理作用中对齐囊泡融合。SNARE与离子通道在动物中的相互作用似乎有利于电信号和神经内分泌分泌，我们已经表明，拟南芥质膜SNARE SYP 121和离子通道亚基KC 1之间的相互作用会影响运输的植物活性K+离子。一个明显的潜在推论是SNARE和离子通道之间的结合在调节囊泡融合中是重要的。尽管这一过程的基本性质，我们对植物或其他有壁真核细胞中细胞体积与渗透溶质运输耦合的分子机制知之甚少。事实上，有壁细胞如何调节与细胞体积平行的抗肿瘤活性溶质（特别是K+）的运输-无论是在保卫细胞中可逆还是在不可逆的扩张生长期间-仍然是一个相当大的争论问题。这一建议建立在我们最近对SYP 121-KC 1蛋白复合物及其在K+通道调节中的作用的鉴定的重要发现基础上：（1）我们已经鉴定了SNARE蛋白SYP 121上结合通道亚基KC 1的位点，（2）我们已经证明KC 1与Sec 1/Munc 18（SM）蛋白KEULE竞争结合SYP 121。SM蛋白是所有真核生物中膜囊泡融合的关键调节因子，但它们在控制SNARE介导的膜融合中的确切作用仍然不清楚，并且是一个深入研究的主题。我们的研究结果指出SM蛋白功能和SNARE通道相互作用之间的联系。他们不仅提供了第一个证据的机制，直接耦合渗透溶质运输与囊泡交通控制细胞扩张壁真核生物，但他们也支持SM调节膜交通的新模型。我们的工作假设是，SYP 121和KC 1的SNARE-K+通道相互作用充当“分子总督”，类似于James Watt在调节他的蒸汽机的周转率时所采用的机械发明，以协调囊泡交通（和细胞扩张）与吸收具有生物活性的K+离子。我们现在打算检验这个假设的各个方面。我们的目标是充分表征SYP 121与KC 1和KEULE的结合，以建立相互作用的分子图谱。我们还建议研究选择性地破坏SYP 121，KC 1和KEULE之间的相互作用对SNARE复合物组装，膜交通和通道介导的K+运输的后果。我们的多学科的方法不仅将进一步我们的渗透溶质运输和控制细胞体积之间的联系的理解，但也有可能提供一个新的范例连接膜交通与其他生理过程。

项目成果

Exploring emergent properties in cellular homeostasis using OnGuard to model K+ and other ion transport in guard cells.

• DOI: 10.1016/j.jplph.2013.09.014
• 发表时间: 2014-05-15
• 期刊: JOURNAL OF PLANT PHYSIOLOGY
• 影响因子: 4.3
• 作者: Blatt, Michael R.;Wang, Yizhou;Leonhardt, Nathalie;Hills, Adrian
• 通讯作者: Hills, Adrian

The conceptual approach to quantitative modeling of guard cells

• DOI: 10.4161/psb.22747
• 发表时间: 2013-01-01
• 期刊: PLANT SIGNALING & BEHAVIOR
• 影响因子: 2.9
• 作者: Blatt, Michael R.;Hills, Adrian;Lew, Vigilio L.
• 通讯作者: Lew, Vigilio L.