Phosphoinositide Domains and Gradients in Asymmetric Lipid Bilayers: Physical Properties and Protein Recognition

不对称脂质双层中的磷酸肌醇结构域和梯度：物理性质和蛋白质识别

• 批准号: 1904886
• 负责人: Arne Gericke
• 金额: $ 50.97万
• 依托单位: Worcester Polytechnic Institute
• 依托单位国家: 美国
• 项目类别: Standard Grant
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-08-15 至 2023-07-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1904886&HistoricalAwards=false
• 关键词: Phosphoinositide; Domains; Gradients; Asymmetric; Lipid

Lipids are a class of biomolecules that affect or even control virtually all processes that occur on cellular membranes. They are very important for cell physiology and human biology. Biological membranes are bilayers of lipids. The two parts of the bilayer have different overall lipid composition as well as different lateral composition. Each part contains domains that have distinct lipid compositions and physical properties. Different proteins are attracted to these domains. As a result, lipids and proteins form supramolecular structures that are platforms for a large number of important cellular functions, most notably cell signaling. With this award, the Chemistry of Life Processes Program in the Chemistry Division and the Molecular Biophysics Program in the Division of Molecular and Cellular Biosciences are funding Dr. Arne Gericke from Worcester Polytechnic Institute to investigate the domains in biological membrane mimics. More specifically, the Gericke research group investigates whether the domains situated in the two opposing bilayers line up ("register") with each other and if so, whether the two domains situated in register physically affect each other. Undergraduate, graduate and postdoctoral students participate in the research. The undergraduate students have the opportunity to participate in a 10-week summer research experience at the Technical University Dortmund, Germany. Dr. Gericke continues to develop and teach a summer camp for middle school students that introduces the students to the unique properties of water as the foundation for all life, to the properties of detergents and lipids, and to the unique and important properties of biological membranes. Phosphoinositide (PIPs) lipids affect an extraordinary variety of protein functions. The specificity of PIP-mediated signaling is rooted in the chemical properties of the lipid's headgroup, which mediates specific interactions with distinct protein motifs. Temporal control of signaling is provided by kinases and phosphatases that phosphorylate or dephosphorylate PIPs. The spatial control of PIP-mediated signaling stems from the non-uniform lateral distribution of PIPs in the cell membrane, which gives rise to PIP-enriched domains and gradients. Rafts, which are liquid-ordered domains enriched in sphingolipid and cholesterol, are found in the outer leaflet of the plasma membrane and affect signaling events on the inner leaflet. Many PIP-mediated signaling events have been associated with lipid rafts; however, given the chemical nature of PIPs, they are expected to partition into fluid membrane patches rather than rafts. Aim 1 addresses this long-standing conundrum by fabricating asymmetric lipid bilayers that exhibit a lipid-raft mixture on the outer leaflet and PIP-containing lipid mixtures on the inner leaflet. Dr. Gericke's group uses these asymmetric lipid bilayers to determine the conditions that result in domain registration. Aim 2 explores how three peripheral membrane proteins, MARCKS, PTEN, and Akt, interact with three types of PIP domains (fluid PIP/cholesterol, liquid-ordered PIP/cholesterol, and PIP/cholesterol in contact with a raft domain in the opposing leaflet). The MARCKS interactions with PIPs are non-specific, purely electrostatic, PTEN binds phosphatidylinositol-4,5-bisphosphate, and the Akt protein has a pocket to which phosphatidylinositol-4,5-bisphosphate binds. Lipid gradients are crucial for important cellular processes, including cytokinesis and directed cell movement. While cell-based studies have highlighted the importance of lipid gradients for cellular function, the physiochemical underpinnings of lipid gradients have not been addressed. Dr. Gericke's group has developed a microfluidic lipid gradient generator to systematically study how lipid gradients interact and/or protein gradients in the aqueous phase above the lipid bilayer create and maintain lipid gradients. In aim 3, Dr. Gericke investigates how opposing gradients of PI3K and PTEN create lipid gradients and the effect of pre-existing domains on gradient development.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.

脂质是一类影响甚至控制几乎所有发生在细胞膜上的过程的生物分子。它们对细胞生理学和人类生物学非常重要。生物膜是脂质双层。双分子层的两个部分具有不同的总体脂质组成以及不同的横向组成。每个部分都包含具有不同脂质组成和物理性质的结构域。不同的蛋白质被吸引到这些结构域。因此，脂质和蛋白质形成的超分子结构是大量重要细胞功能的平台，最显著的是细胞信号传导。有了这个奖项，化学部门的生命过程化学项目和分子和细胞生物科学部的分子生物物理项目资助了伍斯特理工学院的Arne Gericke博士研究生物膜模拟领域。更具体地说，Gericke研究小组调查了位于两个对立双层中的域是否彼此排列（“寄存器”），如果是，位于寄存器中的两个域是否在物理上相互影响。本科生、研究生和博士后参与研究。本科生有机会在德国多特蒙德技术大学参加为期10周的暑期研究体验。Gericke博士继续为中学生开发和教授夏令营，向学生介绍作为所有生命基础的水的独特特性，洗涤剂和脂质的特性，以及生物膜的独特和重要特性。磷脂酰肌苷（PIPs）脂类影响多种蛋白质功能。pip介导的信号传导的特异性植根于脂质头基团的化学性质，它介导与不同蛋白质基序的特异性相互作用。信号传导的时间控制是由磷酸化或去磷酸化pip的激酶和磷酸酶提供的。pip介导的信号的空间控制源于细胞膜中pip的不均匀横向分布，这产生了pip富集区域和梯度。筏是富含鞘脂和胆固醇的液体有序结构域，存在于质膜的外小叶中，影响内小叶的信号事件。许多pip介导的信号事件与脂筏有关；然而，考虑到pip的化学性质，它们预计会分裂成流体膜斑块而不是筏。目的1通过制造不对称的脂质双分子层来解决这个长期存在的难题，这种双分子层在外层小叶上表现为脂质筏混合物，在内层小叶上表现为含有pip的脂质混合物。Gericke博士的研究小组利用这些不对称脂质双分子层来确定导致域注册的条件。目的2探讨了三种外周膜蛋白，MARCKS， PTEN和Akt，如何与三种类型的PIP结构域（液体PIP/胆固醇，液体有序PIP/胆固醇，以及与相对小叶中的筏结构域接触的PIP/胆固醇）相互作用。MARCKS与pip的相互作用是非特异性的，纯静电的，PTEN结合磷脂酰肌醇-4,5-二磷酸，Akt蛋白有一个袋，磷脂酰肌醇-4,5-二磷酸结合。脂质梯度对重要的细胞过程至关重要，包括细胞分裂和定向细胞运动。虽然基于细胞的研究强调了脂质梯度对细胞功能的重要性，但脂质梯度的物理化学基础尚未得到解决。Gericke博士的团队开发了一种微流体脂质梯度发生器，系统地研究脂质梯度如何相互作用和/或脂质双分子层上方水相中的蛋白质梯度如何产生和维持脂质梯度。在目标3中，Gericke博士研究了PI3K和PTEN的相反梯度是如何产生脂质梯度的，以及预先存在的结构域对梯度发展的影响。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

项目成果

IQGAP1 scaffolding links phosphoinositide kinases to cytoskeletal reorganization

• DOI: 10.1016/j.bpj.2022.01.018
• 发表时间: 2022-03-01
• 期刊: BIOPHYSICAL JOURNAL
• 影响因子: 3.4
• 作者: Yerramilli，V. Siddartha;Ross，Alonzo H.;Gericke，Arne
• 通讯作者: Gericke，Arne