Bipartite regulation of cellular osmosensing in C. elegans

描述(申请人提供)：维持细胞溶质和水分含量的生理过程被称为渗透平衡，或渗透调节，对所有形式的细胞生命都是必不可少的。在人类中，渗透平衡在几种情况下发挥着至关重要的作用，包括调节肾脏的尿液浓缩机制，控制血压，以及激活免疫反应。渗透压失调与几种与年龄相关的疾病有关，包括慢性肾脏疾病、肾功能衰竭、高血压和周围神经病。尽管渗透调节在生理和病理生理疾病状态下都很重要，但对动物细胞感知和响应渗透胁迫的机制知之甚少。更好地了解这些机制可能有助于更早地发现和干预与年龄相关的疾病。大多数渗透调节的研究都是使用培养细胞进行的，这无法模拟大多数细胞发现的复杂环境。这些研究导致了许多假说来解释细胞渗透感知的机制(S)，如膜和/或细胞骨架的机械‘拉伸’，大分子拥挤，以及细胞质离子含量的变化，仅举几例。然而，几乎没有数据支持这些模型中的任何一个。为了了解动物体内的细胞渗透感知机制，我们在模式生物秀丽线虫中研究了这一过程，在这个过程中，复杂的细胞-细胞和细胞-细胞外基质(ECM)相互作用被保留。使用无偏见的正向和反向遗传方法，我们发现了细胞外基质(Rohlfing等人，PLoS Genetics，2011)和蛋白质错误折叠(Moronetti Mazzeo等人，PNAS，2012)在线虫细胞渗透感觉调控中的关键作用。基于这些发现，我们假设动物细胞使用机械转导和蛋白质损伤检测机制来感知渗透紊乱并激活渗透敏感基因的表达。在目标1中，我们将确定线虫角质层细胞外基质是否作为结构‘渗透传感器’，通过粘蛋白样蛋白OSM-8和跨膜蛋白PTR-23之间的相互作用来传递信息。在目标2中，我们将定义对应激诱导的蛋白质聚集敏感的天然蛋白质，并确定衰老和衰老调节剂如何影响渗透胁迫诱导的蛋白质损伤和渗透敏感基因的表达。在目标3中，我们将研究细胞外基质和蛋白质损伤检测通路如何相互作用来控制渗透压调节生理。我们的研究最大限度地利用线虫系统来填补我们在后生动物细胞生理学知识方面的一个重要空白。这些发现将为渗透调节的保守过程提供变革性的见解，使我们能够更好地理解、检测和管理与年龄相关的渗透调节失调疾病。

DESCRIPTION (provided by applicant): The physiological process of maintaining cellular solute and water content is termed osmotic homeostasis, or osmoregulation, and is essential for all forms of cellular life. In humans, osmotic homeostasis plays vital roles in several contexts, including regulation of the kidney's urinary concentrating mechanism, control of blood pressure, and activation of immune responses. Osmotic dyshomeostasis is associated with several age- related diseases, including chronic kidney disease, renal failure, hypertension, and peripheral neuropathy. Despite the obvious importance of osmoregulation in both physiological and pathophysiological disease states, little is known about the mechanisms by which animal cells sense and respond to osmotic stress. A better understanding of these mechanisms may allow earlier detection and intervention in age-related diseases. Most studies of osmoregulation have been carried out using cultured cells, which fail to mimic the complex environments in which most cells are found. These studies have led to many hypotheses to explain the mechanism(s) of cellular osmosensing, such as mechanical 'stretching' of the membrane and/or cytoskeleton, macromolecular crowding, and alterations in cytoplasmic ionic content, to name a few. However, there is little data supporting any of these models. To gain an in vivo perspective on mechanisms of cellular osmosensing in animals, we are studying this process in the model organism C. elegans, in which complex cell-cell and cell- extracellular matrix (ECM) interactions are preserved. Using unbiased forward and reverse genetic approaches, we discovered critical roles for the extracellular matrix (Rohlfing et al, PLoS Genetics, 2011) and protein misfolding (Moronetti Mazzeo et al, PNAS, 2012) in the regulation of cellular osmosensing in C. elegans. Based on these findings we hypothesize that animal cells use both mechanotransduction and protein damage detection mechanisms to sense osmotic disturbances and activate osmosensitive gene expression. In Aim 1, we will determine if the C. elegans cuticular ECM acts as a structural 'osmosensor' to transduce information via interactions between the mucin-like protein OSM-8 and a transmembrane protein PTR-23. In Aim 2, we will define the native proteins susceptible to stress-induced protein aggregation and determine how aging and aging regulators influence osmotic stress induced protein damage and osmosensitive gene expression. In Aim 3, we will examine how ECM and protein damage detection pathways interact with each other to control osmoregulatory physiology. Our studies take maximal advantage of the C. elegans system to fill an important gap in our knowledge of metazoan cell physiology. These findings will provide transformative insights into the conserved process of osmoregulation that will allow us to better understand, detect, and manage age-related diseases of osmotic dyshomeostasis.