NHE3 Polymorphisms and Chronic Diarrheal Diseases

NHE3 多态性与慢性腹泻病

• 批准号: 7953165
• 负责人: Xinjun Zhu
• 金额: $ 13.8万
• 依托单位: ALBANY MEDICAL COLLEGE
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-09-01 至 2015-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7953165
• 关键词: Accounting; Acute; Address; Award; Binding Proteins; Bioinformatics; Biology; Biomedical Research; Biophotonics; Biotin; Brain; C-terminal; Calcineurin; Calcium; Chromatography; Chronic; Co-Immunoprecipitations; Colon; Complex; Confocal Microscopy; Cytoplasmic Tail; Data; Defect; Dexamethasone; Diarrhea; Disease; Education; Energy Transfer; Enrollment; Environment; Exhibits; Failure; Fostering; Foundations; Functional disorder; Funding; Gastroenterology; Genes; Genetic; Genetic Determinism; Genetic Polymorphism; Goals; Grant; Homeostasis; Homologous Protein; Hydrogen; Inherited; Kidney; Knockout Mice; Knowledge; Label; Life; Location; Mass Spectrum Analysis; Medicine; Mentors; Molecular; Monitor; Mutation; Patients; Phenotype; Photobleaching; Principal Investigator; Program Development; Proteins; Proteomics; Regulation; Research; Research Personnel; Role; Schools; Scientist; Single Nucleotide Polymorphism; Site; Small Intestines; Sodium; Solid; Streptavidin; Surface; System; Training; Training Programs; Transferase; Universities; Water; absorption; base; career; cell fixing; experience; ezrin; genetic regulatory protein; in vivo; insight; mutant; new therapeutic target; novel; programs; protein complex; protein profiling; protein protein interaction; public health relevance; research study; response; skills; sodium-hydrogen exchanger 3; trafficking

DESCRIPTION (provided by applicant): This proposal describes a 5-year training program for the development of an academic research career in Gastroenterology and Medicine. The proposal builds upon the candidate's strengths and prior research skills, and takes advantage of a vibrant and stimulating scientific environment at The Johns Hopkins University. The principal investigator will be supported and mentored by Dr. Mark Donowitz, a leader in Na/H exchanger biology and diarrheal diseases. In addition, a panel of accomplished and well-established scientists will regularly provide the principal investigator with scientific and career advice. The goal is to foster the PI to develop a successful, independently funded research program and to gain the experience necessary to maintain a productive scientific career relevant to research on diarrheal disease. The objective of the proposed research is to identify and characterize the molecular and cellular mechanisms responsible for the dysfunction of sodium/hydrogen exchanger 3 (NHE3) we identified are present in NHE3 polymorphisms. Previous studies revealed that diarrhea is the major phenotype in NHE3 knockout mice, suggesting a potential involvement of NHE3 in diarrheal diseases. Single nucleotide polymorphisms (SNPs) account for many inherited diseases. Our preliminary studies 1) identified three missense SNPs in the NHE3 cytoplasmic tail, the regulatory domain for NHE3; 2) demonstrated that these mutants, R474Q, V567M and R799C, caused reduced basal NHE3 activity; 3) revealed the abnormal function of these SNPs in NHE3 appears to be due to either a change in intrinsic NHE3 function in R474Q, or abnormal trafficking in V567M and defect in both intrinsic NHE3 function and trafficking in R799C; 4) demonstrated that R474Q and R799C mutants failed to respond to acute dexamethasone stimulation, suggesting a possible defect of NHE3 in response to stimulation postprandially; and 5) identified a mutation of NHE3 at a polymorphic site R474K in 2 of 3 patients with chronic diarrhea due to congenital sodium diarrhea (CSD) and this mutant failed to respond to dexamethasone. These findings suggest that poorly functioning NHE3 polymorphisms may act as genetic determinants that contribute to chronic diarrheal diseases. However, the molecular mechanisms by which the identified three NHE3 polymorphisms cause reduced sodium absorption remain ill defined. In this proposal, we will determine whether NHE3 protein complex size is altered in polymorphisms under basal and dexamethasone-stimulated conditions. We will also investigate whether the interactions of NHE3 mutants and NHE3 known regulatory proteins are changed under basal and dexamethesone- stimulated conditions. Finally, we will use proteomic approaches to identify novel proteins associated with NHE3 polymorphisms. Together, experiments proposed in these three specific aims will provide insight into the genetic basis of chronic diarrheal disease with the ultimate goal of establishing NHE3 and related proteins as novel therapeutic targets for treatment of these diarrheal diseases. This grant will also allow me to develop a solid foundation of knowledge in chronic diarrheal diseases and the qualifications of an independent investigator. To achieve this goal, formal education during the award period will be obtained through enrolling and auditing courses offered by the JHU Graduate School. Specific courses that are relevant to my training, but not limited to, are: BIophotonics (585.634) and Fundamentals of confocal Microscopy (110.807), Bioinformatics (410.639) and Role of chromatography and Mass Spectrometry in Biomedical research (330.804). PUBLIC HEALTH RELEVANCE: Sodium and hydrogen absorption that occurs in small intestine and colon is mainly by regulation of NHE3 activity. Inhibition of NHE3 activity is present in polymorphic NHE3. However, the mechanisms responsible for these poorly functioning NHE3 polymorphisms remain unknown. The current study will provide novel insight s into understanding genetic component accounts for the role of NHE3 in chronic diarrheal diseases.

描述（由申请人提供）： 该提案描述了一个为期5年的培训计划，用于发展胃肠病学和医学的学术研究事业。该提案基于候选人的优势和先前的研究技能，并利用约翰霍普金斯大学充满活力和刺激的科学环境。首席研究员将得到Na/H交换生物学和牙周病领域的领导者Mark Donowitz博士的支持和指导。此外，一个由有成就和有声望的科学家组成的小组将定期向主要研究员提供科学和职业建议。其目标是促进PI制定一个成功的，独立资助的研究计划，并获得必要的经验，以保持与疟疾研究相关的富有成效的科学事业。拟议研究的目的是确定和表征负责钠/氢交换器3（NHE 3）功能障碍的分子和细胞机制，我们确定NHE 3多态性存在。先前的研究表明，腹泻是NHE 3基因敲除小鼠的主要表型，这表明NHE 3可能参与了肠道疾病。单核苷酸多态性（SNP）是许多遗传性疾病的原因。我们的初步研究1）在NHE 3胞质尾区（NHE 3的调控结构域）鉴定了三个错义SNP：2）证明这些突变体R474 Q、V567 M和R799 C导致NHE 3基础活性降低; 3）揭示了NHE 3中这些SNP的异常功能似乎是由于R474 Q中内在NHE 3功能的改变，或V567 M异常运输和内在NHE 3功能和R799 C运输缺陷; 4）证明R474 Q和R799 C突变体不能对急性地塞米松刺激应答，表明NHE 3在对餐后刺激应答中可能存在缺陷;（5）在3例先天性钠性腹泻（CSD）慢性腹泻患者中发现2例NHE 3基因R474 K多态性位点突变，该突变对地塞米松无效。这些发现表明，功能不良的NHE 3多态性可能作为遗传决定因素，有助于慢性牙周病。然而，确定的三个NHE 3多态性导致钠吸收减少的分子机制仍然不清楚。在这个建议中，我们将确定是否NHE 3蛋白复合物的大小改变基础和地塞米松刺激条件下的多态性。我们还将研究NHE 3突变体和NHE 3已知调节蛋白的相互作用是否在基础和地塞米松刺激条件下改变。最后，我们将使用蛋白质组学方法来鉴定与NHE 3多态性相关的新蛋白质。总之，在这三个具体目标中提出的实验将提供对慢性牙周炎疾病的遗传基础的深入了解，最终目标是建立NHE 3和相关蛋白作为治疗这些牙周炎疾病的新的治疗靶点。 这笔赠款还将使我能够在慢性牙周病方面建立坚实的知识基础，并获得独立调查员的资格。为了实现这一目标，在奖励期间的正规教育将通过注册和审计JHU研究生院提供的课程获得。与我的培训相关的具体课程，但不限于：生物光子学（585.634）和共聚焦显微镜基础（110.807），生物信息学（410.639）和色谱和质谱在生物医学研究中的作用（330.804）。 公共卫生关系： 在小肠和结肠中发生的钠和氢吸收主要通过调节NHE 3活性。NHE 3活性的抑制存在于多态性NHE 3中。然而，负责这些功能不良的NHE 3多态性的机制仍然未知。目前的研究将为理解NHE 3在慢性牙周病中的作用提供新的见解。