Regulation of epithelial cell homeostasis by actin microfilaments

肌动蛋白微丝对上皮细胞稳态的调节

基本信息

批准号：
7746802
负责人：
Seema Khurana
金额：
$ 37.23万
依托单位：
UNIVERSITY OF TENNESSEE HEALTH SCI CTR
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-08-20 至 2011-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7746802
关键词：
Actin-Binding Protein Actins Affect Apoptosis Apoptotic Architecture Atrophic Biochemical Celiac Disease Cell Survival Cells Cleaved cell Colon Carcinoma Colorectal Cancer Communicable Diseases Complex Data Defect Developed Countries Developing Countries Disease Drug usage Epithelial Epithelial Cells Equilibrium Eukaryotic Cell Evaluation Exocrine Glands Family Funding Gastrointestinal tract structure Gelsolin Healed Homeostasis Homologous Protein Hyperplasia Immune Infection Inflammatory Inflammatory Bowel Diseases Intervention Intestines Ischemia Knockout Mice Large Intestine Length Link Location Maintenance Malignant Neoplasms Microfilaments Mitochondria Mitochondrial Proteins Molecular Morphology Mus N-terminal Organ Parasitic infection Pathogenesis Persons Pharmaceutical Preparations Phenotype Physiological Physiology Play Protein Family Proteins Public Health Radiation therapy Regulation Reperfusion Injury Reporting Research Resistance Risk Role Stimulus Surface Time Villus Viral Yeasts base biological adaptation to stress cancer cell chemotherapy design economic impact effective therapy gastrointestinal gastrointestinal epithelium genetic regulatory protein healing in vivo intestinal epithelium novel prevent public health relevance research study social tumorigenesis villin yeast two hybrid system

项目摘要

DESCRIPTION (provided by applicant): The homeostatic balance between proliferation and apoptosis is essential for the intestinal epithelium to function as a physiological and structural barrier. Intestinal epithelial cells have a high rate of cell turnover accompanied by an equally high rate of apoptosis. This normal apoptosis is essential for the hierarchical organization of the intestinal epithelium and apoptotic epithelial cells have been detected both at the base of the crypt as well as the villus tips of the small and large intestine. Defects in apoptosis are associated with several gastrointestinal conditions including villus atrophy, epithelial hyperplasia, loss of normal absorptive function and increased risk of tumorigenesis. Surprisingly, there are few data available on apoptosis as an important mechanism of action in vivo in the gastrointestinal epithelium. There are very comprehensible data supporting links between actin and apoptosis in eukaryotic cells from studies using drugs that affect actin turnover. However, few studies have established a causal link between microfilament reorganization and cell survival. Villin is an actin regulatory protein expressed in the gastrointestinal epithelial cells of the small and large intestine as well as in exocrine glands associated with the GI tract. We recently determined, for the first time, that villin is an epithelial cell-specific anti-apoptotic protein. Further, our preliminary studies suggest that in the absence of villin, this function may be fulfilled by a related actin- binding protein of the villin family, gelsolin. Villin is also cleaved during the apoptotic cycle and we have previously reported that the cleaved NH2-terminal fragment of villin is pro-apoptotic. Using yeast two hybrid, biochemical and cellular approaches, we have recently identified and validated the novel interaction of villin with the mitochondrial protein, ATP synthase 2-subunit, which now allows us to investigate the cellular and molecular mechanism(s) of microfilament- induced regulation of mitochondrial morphology and function and epithelial cell survival by villin. The novelty of our observation will allow us to demonstrate for the first time a causal relationship between microfilament regulation and cell survival in eukaryotic cells. To achieve this, experiments will be conducted under three major specific aims: (i) to characterize the cellular and molecular mechanism(s) of villin's anti- and pro-apoptotic functions and to identify the physiologic relevance of full-length and cleaved villin fragments respectively, in the maintenance of gastrointestinal homeostasis ; (ii) to characterize the interaction of villin with ATP synthase 2- subunit to examine the relevance of this complex in the maintenance of gastrointestinal homeostasis; (iii) to characterize the villin-gelsolin double knock out mice to establish the role of villin and its homologous protein gelsolin in the regulation of gastrointestinal homeostasis. Our studies are vital for the rational design of new and more effective therapies for major intestinal epithelial diseases such as inflammatory bowel disease and cancer. PUBLIC HEALTH RELEVANCE: The intestinal tract fulfils essential roles as a digestive and absorptive surface and constitutes the largest immune organ. In addition the intestine is the anatomical location of diseases of enormous social and economic impact, like the inflammatory bowel disease (affecting about 1 person in every 500 in USA), the colon carcinoma (the second most frequent malignancy in developed countries) and numerous infectious diseases. Apoptosis plays an important role in determining the architecture of intestinal epithelia and also a part of the stress response of intestinal epithelial cells to toxic stimuli. In disease pathogenesis, apoptosis can either be inappropriately excessive or deficient and both of these have been implicated in bacterial, viral and parasitic infections, ischemia-reperfusion injury, inflammatory bowel disease, celiac disease and colorectal cancer. Abnormalities in apoptotic function contribute to both the pathogenesis of colorectal cancer as well as its resistance to chemotherapeutic drugs and radiotherapy. An understanding of apoptosis would help enhance normal physiology and ameliorate diseases. Example of such intervention include hastening the healing or preventing chemotherapy- and radiotherapy-induced intestinal damage; to overcome apoptosis resistance in cancer cells; to retard the damage induced in inflammatory conditions such as inflammatory bowel disease, celiac disease, enteropathogenic infections and ischemia.

描述（由申请人提供）：增殖和凋亡之间的稳态平衡对于肠上皮作为生理和结构屏障的功能至关重要。肠上皮细胞具有高细胞更新率，同时伴随着同样高的细胞凋亡率。这种正常的细胞凋亡对于肠上皮的分层组织至关重要，并且在隐窝底部以及小肠和大肠的绒毛尖端都检测到了凋亡的上皮细胞。细胞凋亡缺陷与多种胃肠道疾病有关，包括绒毛萎缩、上皮增生、正常吸收功能丧失和肿瘤发生风险增加。令人惊讶的是，关于细胞凋亡作为胃肠道上皮体内重要作用机制的数据很少。使用影响肌动蛋白周转的药物进行的研究中有非常容易理解的数据支持肌动蛋白与真核细胞凋亡之间的联系。然而，很少有研究确定微丝重组与细胞存活之间的因果关系。绒毛蛋白是一种肌动蛋白调节蛋白，在小肠和大肠的胃肠上皮细胞以及与胃肠道相关的外分泌腺中表达。我们最近首次确定绒毛蛋白是一种上皮细胞特异性抗凋亡蛋白。此外，我们的初步研究表明，在没有绒毛蛋白的情况下，这一功能可能由绒毛蛋白家族的相关肌动蛋白结合蛋白凝溶胶蛋白来实现。绒毛蛋白也在细胞凋亡周期中被裂解，我们之前报道过绒毛蛋白的 NH2 末端裂解片段是促凋亡的。使用酵母两种杂交、生化和细胞方法，我们最近鉴定并验证了绒毛蛋白与线粒体蛋白、ATP 合酶 2 亚基的新型相互作用，这使我们能够研究微丝诱导的细胞和分子机制。绒毛蛋白对线粒体形态和功能以及上皮细胞存活的调节。我们观察的新颖性将使我们能够首次证明真核细胞中微丝调节与细胞存活之间的因果关系。为了实现这一目标，实验将在三个主要具体目标下进行：（i）表征绒毛蛋白抗和促凋亡功能的细胞和分子机制，并确定全长和切割绒毛片段的生理相关性分别维持胃肠道稳态； (ii) 表征绒毛蛋白与 ATP 合酶 2-亚基的相互作用，以检查该复合物在维持胃肠道稳态中的相关性； (iii)表征绒毛蛋白-凝溶胶蛋白双敲除小鼠，以确定绒毛蛋白及其同源蛋白凝溶胶蛋白在调节胃肠道稳态中的作用。我们的研究对于合理设计针对炎症性肠病和癌症等主要肠上皮疾病的新的、更有效的疗法至关重要。公共卫生相关性：肠道作为消化和吸收表面发挥着重要作用，并构成最大的免疫器官。此外，肠道是具有巨大社会和经济影响的疾病的解剖部位，例如炎症性肠病（美国每 500 人中约有 1 人受影响）、结肠癌（发达国家第二常见的恶性肿瘤）和许多传染性肠病。疾病。细胞凋亡在决定肠上皮细胞结构中起重要作用，也是肠上皮细胞对毒性刺激应激反应的一部分。在疾病发病机制中，细胞凋亡可能过度或不足，这两种情况都与细菌、病毒和寄生虫感染、缺血再灌注损伤、炎症性肠病、乳糜泻和结直肠癌有关。细胞凋亡功能的异常有助于结直肠癌的发病机制及其对化疗药物和放射治疗的抵抗。了解细胞凋亡将有助于增强正常生理机能并改善疾病。这种干预的例子包括加速愈合或预防化疗和放疗引起的肠道损伤；克服癌细胞的凋亡抵抗；延缓炎症性疾病（如炎症性肠病、乳糜泻、肠道病原性感染和缺血）引起的损害。