CHD1, Chromatin Dynamics, and Salivary Gland Differentiation

描述（申请人提供）：唾液腺的分化和功能依赖于唾液的产生和分泌。黏液主要由舌下腺（SLGs）产生，赋予唾液粘稠和防腐的特性。粘蛋白糖蛋白是粘膜的主要成分，是形成粘膜屏障所必需的。slg分泌的粘蛋白减少导致口干症，其特征是发音和吞咽困难，粘膜溃疡和龋齿的频率增加。口干症的主要原因之一是一种被称为干燥综合征的自身免疫性疾病，它会影响人体的多个器官，最明显的是唾液腺和泪腺。fs - sid小鼠，干燥综合征模型，表现出SLG分化延迟，表现为新生儿粘蛋白缺乏，特别是小鼠SLG的主要粘蛋白产物Muc19。有趣的是，我们的实验室已经证实了染色质重塑蛋白CHD1在调节Muc19表达中的作用。该项目旨在进一步研究通过CHD1及其相互作用物范围内slg控制粘蛋白产生的分子机制，并将这些发现与开发重新激活粘蛋白产生的新治疗靶点联系起来。目的1：比较NFS-sId和NFS-N（对照）小鼠中CHD1和Mud9的表达水平、CHD1蛋白水平以及CHD1在Muc19启动子和转录区域的物理存在。目的2：探讨CHD1调控Muc19基因的方法发生在转录延伸和/或选择性剪接水平的可能性。所有实验均使用NFS-sId和对照NFS-N小鼠。基因表达和免疫印迹分析，结合体内免疫沉淀实验，将揭示CHD1及其相互作用物的数量、定位和伙伴关系对Muc19表达和随后产生的影响，而额外的体内dna -蛋白质相互作用研究，结合Muc19转录变异体的研究，将有助于推断Muc19基因在SLGs中调控的具体机制。在SLG中控制粘蛋白产生的基因调控机制仍然知之甚少。通过完成该项目获得的知识将加强对粘蛋白分泌减少相关的流行病学的理解，例如发生在干燥综合征患者身上的粘蛋白分泌减少。此外，可以确定旨在重新激活SLG功能的新治疗靶点。

DESCRIPTION (provided by applicant): Salivary gland differentiation and functionality relies on the production and secretion of saliva. Mucous, produced mainly by the sublingual glands (SLGs), confers viscous and antiseptic properties on saliva. Mucin glycoproteins are the major constituent of mucous, essential for the formation of mucosal barriers. Hyposecretion of mucins by the SLGs results in xerostomia, characterized by difficulty in phonation and deglutition, mucosal ulcerations, and an increase in the frequency of dental caries. One of the major contributors to xerostomia is an autoimmune disorder known as Sjogren's Syndrome which affects multiple body organs, most notably the salivary and lacrimal glands. NFS-sId mice, a model for Sjogren's Syndrome, display delayed SLG differentiation noted by neonatal mucin-deficiencies, specifically the major mucin product of mouse SLGs, Muc19. Interestingly, our laboratory has confirmed a function for the chromatin- remodeling protein CHD1 in regulating Muc19 expression. This project is designed to further investigate the molecular mechanisms that govern mucin production by the SLGs through the scope of CHD1 and its interactors, relating these findings to the development of novel therapeutic targets for re-activation of mucin production. To accomplish this, the following specific aims will be addressed: Aim 1: Compare levels of CHD1 and Mud9 expression, CHD1 protein levels, and the physical presence of CHD1 on the Muc19 promoter and transcribed regions, in NFS-sId and NFS-N (control) mice. Aim 2: Investigate the possibilities that the method of Muc19 gene regulation by CHD1 is occurring at the transcriptional elongation and/or alternative-splicing level. NFS-sId and control NFS-N mice will be utilized for all experiments. Gene expression and immunoblotting assays, combined with in vivo immunoprecipitation experiments, will shed light on the influence of quantity, localization, and partnership of CHD1 and its interactors on the expression, and subsequent production, of Muc19, while additional in vivo DNA-protein interaction studies, combined with an investigation of Muc19 transcript variants will allow inferences on the specific mechanism of Muc19 gene regulation in SLGs. The gene-regulatory mechanisms that control mucin production in the SLG remain poorly understood. Knowledge gained through the completion of this project will enhance the understanding of the epidemiology related to the hyposecretion of mucins, such as that which occurs in those afflicted with Sjogren's Syndrome. Also, novel therapeutic targets aimed at re-activation of SLG functionality could be identified.