The mechanism of cell size regulation by polycystins

多囊蛋白调节细胞大小的机制

基本信息

批准号：
10609393
负责人：
Qian Chen
金额：
$ 31.34万
依托单位：
UNIVERSITY OF TOLEDO
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-04-15 至 2027-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10609393
关键词：
Actin-Binding Protein Actins Actomyosin Autosomal Dominant Polycystic Kidney Biomass Calcineurin Calcium Calcium Signaling Calcium Spikes Calmodulin Cell Cycle Cell Proliferation Cell Separation Cell Size Cell division Cell membrane Cell physiology Cells Collaborations Cytokinesis Cytoskeleton Endocytosis Environment Evolution Family Fission Yeast Genes Genetic Diseases Genetic Screening Growth Human Human Genetics Image In Vitro Intracellular Transport Ion Channel Ions Kidney Diseases Lead Lipids Mediating Michigan Microfilaments Microfluidics Modeling Molecular Morphogenesis Mutation Myosin Type I Myosin Type V Osmosis Osmotic Pressure Pathway interactions Pattern Permeability Phosphotransferases Play Polycystic Kidney Diseases Process Quantitative Microscopy Regulation Regulation of Cell Size Reproducibility Role Signal Pathway Signaling Molecule Stimulus Structure Techniques Tennessee Testing Universities Work Yeasts autosome cell growth constriction daughter cell driving force imaging modality innovation loss of function mutation mechanical force mechanical signal mechanotransduction model organism multidisciplinary novel patch clamp permissiveness public health relevance reconstitution response synergism

项目摘要

The mechanism of mechanosensing by polycystins during cell growth Cytokinesis is the last stage of cell division when two daughter cells separate, but it is equally important for the transition to cell growth including cell size expansion. The mechanism regulating such a transition is poorly understood. We identified the role of calcium and the polycystin channel Pkd2p in this process while studying cytokinesis of the model organism fission yeast. Pkd2p is essential to regulate the cell size and it mediates calcium influx. Polycystins are evolutionally conserved ion channels. Loss of function mutations of human polycystins lead to the genetic disorder, Autosomal Polycystic Kidney Disorder (ADPKD). The cellular function of this highly conserved family of channels is not well understood. This study will determine how Pkd2p and calcium regulate the transition to cell growth. Aim 1. Determine how the Hippo signaling pathways regulate Pkd2p. Our genetic screen has identified the fission yeast Hippo pathways SIN and MOR as crucial for the regulation of Pkd2p. Both are highly conserved kinase cascades that are essential for cell proliferation. SIN antagonizes Pkd2p activity, while MOR synergizes with Pkd2p. However, the mechanism is unclear. We will 1) Determine how MOR and SIN regulate cellular calcium during cytokinesis through calcium-imaging. 2) Determine how MOR and SIN regulate Pkd2p in cell size expansion. 3) Determine how MOR promotes the cell cycle-dependent localization of Pkd2p. 4) Determine whether Pkd2p is a direct substrate of the MOR kinase Orb6p. Aim 2. Determine how the Pkd2p channel is activated. Pkd2 channel opens in response to mechanical force in vitro, but the mechanism is unclear. We will test the proposal that Pkd2p is a channel sensitive to the force driving the yeast cell growth. We will 1) Determine how Pkd2p regulates calcium when the cells are stimulated by external force. 2) Probe how Pkd2p senses osmotic stimuli and the lipid environment in vitro through a collaboration with Allen Liu’s group (University of Michigan). 3) Determine whether Pkd2p channel allows ions other than calcium to pass through, using patch clamp, through a collaboration with Du Jianyang’s group (University of Tennessee). Aim 3 Determine how Pkd2p regulates the actin re-organization during cytokinesis. Calcium can activate two highly conserved molecules Cam1p and Ppb1p. We will determine how they contribute to the role of Pkd2p in cytokinesis. We will determine 1) how Pkd2p regulates the Cam1p-dependent endocytosis during cytokinesis. 2) how Pkd2p regulates the Cam1p-dependent intracellular transport. 3) how Pkd2p regulates the enzymatic activity of Ppb1p. Through this study, we expect to uncover a novel cell size regulation pathway mediated by Pkd2p channel. We will demonstrate how both internal signaling pathways and external environment play a vital role in activating this channel in cytokinesis. We will employ novel imaging methods combined with innovative in vitro techniques in our study. Our works shall help us better understand the cellular functions of the human polycystins.

细胞生长过程中多囊蛋白的机械传感机制细胞分裂是两个子细胞分离时细胞分裂的最后阶段，但它对于细胞分裂同样重要向细胞生长的转变，包括细胞尺寸的扩大，调节这种转变的机制很差。我们在研究过程中确定了钙和多囊蛋白通道 Pkd2p 在此过程中的作用。模型生物体裂殖酵母的胞质分裂对于调节细胞大小至关重要并介导。钙流入。多囊蛋白是人类进化上保守的离子通道。多囊蛋白会导致遗传性疾病，常染色体多囊肾病 (ADPKD)。这个高度保守的通道家族的机制尚不清楚，这项研究将如何确定 Pkd2p 和 Pkd2p 的作用。钙调节细胞生长的过渡。目标 1. 确定 Hippo 信号通路如何调节 Pkd2p。裂殖酵母 Hippo 途径 SIN 和 MOR 对于 Pkd2p 的调节至关重要两者都高度保守。 SIN 拮抗 Pkd2p 活性，而 MOR 则发挥协同作用。然而，其机制尚不清楚 1) 确定 MOR 和 SIN 如何调节细胞。 2) 确定 MOR 和 SIN 如何调节 Pkd2p 的细胞大小 3) 确定 MOR 如何促进 Pkd2p 的细胞周期依赖性定位。 Pkd2p 是否是 MOR 激酶 Orb6p 的直接底物目标 2. 确定 Pkd2p 通道的情况。 Pkd2 通道在体外响应机械力而打开，但其机制尚不清楚。测试 Pkd2p 是对驱动酵母细胞生长的力敏感的通道这一提议，我们将 1) 确定。当细胞受到外力刺激时，Pkd2p 如何调节钙 2) 探究 Pkd2p 如何感知。通过与 Allen Liu 团队（University of University of Technology）合作，研究了渗透刺激和体外脂质环境 3）确定Pkd2p通道是否允许钙以外的离子通过，使用patch 夹子，通过与杜建阳团队（田纳西大学）合作实现目标 3 确定如何进行。 Pkd2p 在胞质分裂过程中调节肌动蛋白重组，钙可以激活两个高度保守的蛋白。我们将确定它们如何促进 Pkd2p 在胞质分裂中的作用。确定 1) Pkd2p 如何在胞质分裂过程中调节 Camp1p 依赖性内吞作用 2) Pkd2p 如何调节。调节 Cam1p 依赖性细胞内运输 3) Pkd2p 如何调节 Ppb1p 的酶活性。通过这项研究，我们期望发现一种由 Pkd2p 通道介导的新型细胞大小调节途径。将展示内部信号通路和外部环境如何在激活中发挥重要作用我们将采用新颖的成像方法与创新的体外技术相结合。在我们的研究中，我们的工作将帮助我们更好地了解人类多囊蛋白的细胞功能。