Functional Analysis of the Bifunctional Ion Channel and Kinase TRPM7

双功能离子通道和激酶 TRPM7 的功能分析

基本信息

批准号：
8601100
负责人：
LOREN W RUNNELS
金额：
$ 33.9万
依托单位：
RBHS-ROBERT WOOD JOHNSON MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-05-01 至 2016-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8601100
关键词：
Adult Affect Biochemical Biological Brain Ischemia Cations Cell Proliferation Cells Congenital Abnormality Defect Development Developmental Gene Embryo Embryonic Development Figs - dietary Funding Homeostasis Homologous Gene Human Incubated Individual Intake Ion Channel Lead Life Magnesium Malignant Neoplasms Mediating Medical Molecular Biology Movement Mus Neural Fold Neural Tube Closure Neural Tube Defects Neural tube Pathway interactions Phosphotransferases Play Pregnancy Prevention strategy Process Proteins Regulation Reporting Research Risk Role Signal Transduction Spinal Dysraphism Staging Stroke Supplementation System Time Tissues United States Xenopus Xenopus laevis Yeasts cancer cell cell behavior cell motility combat cost gain of function gastrulation human disease in vivo innovation insight loss of function mRNA Expression neuron loss novel offspring prevent protein expression protein function public health relevance research study tissue/cell culture xenopus development yeast two hybrid system

项目摘要

DESCRIPTION (provided by applicant): Neural fold closure defect (NTD) is one of the most common birth defects in humans, occurring at an average rate of 1 per 1000 pregnancies. Decreased maternal Mg2+ intake has been associated with an increased risk for NTD, suggesting a key role for Mg2+-permeant ion channels in this essential stage of development. Our research in Xenopus laevis has uncovered important roles for the TRPM6 and TRPM7 ion channels in gastrulation and neural fold closure during embryogenesis. Neural fold closure defects caused by depletion of TRPM7 from Xenopus laevis embryos can be prevented by Mg2+ supplementation or by expression of a Mg2+ transporter, supporting the hypothesis that Mg2+ and the ion channels that conduct this important cation play a critical role during this essential embryonic process. TRPM7 and TRPM6 are known to hetero-oligomerize when heterologously expressed in tissue culture cells, but reports vary as to whether TRPM6 functions by itself as a channel in vivo. Preliminary studies indicate that TRPM6 mRNA expression is upregulated during gastrulation and peaks during neurulation, supporting the hypothesis that the two channels are functioning together to regulate neural fold closure. We propose three specific aims to clarify the function and regulation of these two channels during early development. In the first specific aim, we will employ loss-of-function and gain-of-function experiments in Xenopus laevis to define the role of TRPM6 during development and its connection to the non-canonical Wnt pathway, which has been shown to regulate convergent extension movements during gastrulation and neural fold closure. In specific aim 2 we will examine in Xenopus how TRPM6 and TRPM7 and its individual domains may be functioning together to regulate neural fold closure and how these channels may be impacting Mg2+ homeostasis in the developing embryo. Our research will also focus on how TRPM7's control of Mg2+ homeostasis is affecting the migratory behavior of cells. In specific aim 3 we will investigate the role of 80K-H, a TRPM6- and TRPM7-interacting protein that functions synergistically with TRPM7 during gastrulation and neural fold closure, has in regulating these channels' protein levels, and determine how the Wnt pathway may be impacting this regulation. Collectively, the proposed experiments should greatly advance our understanding how these unique bifunctional channels are functioning in vivo, which could lead to new strategies for preventing neural tube closure defects as well as to new insights for combating the other pathological conditions for which these channels have been associated, including stroke and cancer.

描述（由申请人提供）：神经皱襞闭合缺陷 (NTD) 是人类最常见的出生缺陷之一，平均每 1000 名妊娠中就有 1 人发生这种缺陷。母亲 Mg2+ 摄入量减少与 NTD 风险增加相关，这表明 Mg2+ 渗透离子通道在这一重要的发育阶段发挥着关键作用。我们对非洲爪蟾的研究揭示了 TRPM6 和 TRPM7 离子通道在胚胎发生过程中原肠胚形成和神经折叠闭合中的重要作用。非洲爪蟾胚胎 TRPM7 缺失引起的神经折叠闭合缺陷可以通过补充 Mg2+ 或表达 Mg2+ 转运蛋白来预防，这支持了以下假设：Mg2+ 和传导这种重要阳离子的离子通道在这一重要的胚胎过程中发挥着关键作用。。已知 TRPM7 和 TRPM6 在组织培养细胞中异源表达时会异源寡聚化，但关于 TRPM6 本身是否作为体内通道发挥作用的报道各不相同。初步研究表明，TRPM6 mRNA 表达在原肠胚形成期间上调，并在神经形成期间达到峰值，支持这两个通道共同发挥作用以调节神经折叠闭合的假设。我们提出了三个具体目标，以明确这两个渠道在早期发展过程中的功能和监管。在第一个具体目标中，我们将在非洲爪蟾中进行功能丧失和功能获得实验，以确定 TRPM6 在发育过程中的作用及其与非经典 Wnt 通路的联系，该通路已被证明可以调节收敛原肠胚形成和神经皱襞闭合期间的伸展运动。在具体目标 2 中，我们将在非洲爪蟾中研究 TRPM6 和 TRPM7 及其各个结构域如何共同发挥作用来调节神经折叠闭合，以及这些通道如何影响发育中胚胎中的 Mg2+ 稳态。我们的研究还将重点关注 TRPM7 对 Mg2+ 稳态的控制如何影响细胞的迁移行为。在具体目标 3 中，我们将研究 80K-H 的作用，80K-H 是一种 TRPM6 和 TRPM7 相互作用蛋白，在原肠胚形成和神经折叠闭合过程中与 TRPM7 协同发挥作用，调节这些通道的蛋白水平，并确定 Wnt 通路如何发挥作用会影响该规定。总的来说，所提出的实验应该极大地促进我们对这些独特的双功能通道如何在体内发挥作用的理解，这可能会带来预防神经管闭合缺陷的新策略，以及对抗这些通道所针对的其他病理状况的新见解。相关的，包括中风和癌症。