HIF Dependent Epigenetic Responses During Hypoxia and Differentiation

描述(由申请人提供)： 导师临床科学家发展奖(K08)将允许候选人发展成为一名独立的研究人员，他将在新生儿医学领域进行基础和翻译研究。在这个奖项的过程中，马尔特佩博士将开始一项教学和研究计划，旨在为他提供蛋白质组学和质谱学技术方面的额外培训。他将利用导师在这一领域的专业知识，进一步探索使缺氧诱导因子(HIF)蛋白家族能够调节缺氧反应并决定干细胞命运的表观遗传学机制。具体目标如下。(目的1)分析野生型和HIF缺失型小鼠胚胎成纤维细胞在低氧培养下以及在野生型和HIF缺失型滋养层干细胞分化过程中组蛋白修饰的变化。(目的2)利用免疫沉淀、候选蛋白免疫检测和质谱技术，确定在上述条件下HIF相互作用的染色质重塑复合体的性质。(目的3)利用定量蛋白质组学和质谱学研究上述条件下组蛋白翻译后修饰的整体变化。(目的4)利用BAG克隆阵列和甲基化敏感的限制性内切酶Not1，分析上述条件下全基因组CpG岛甲基化状态的变化。从这些研究中获得的信息将为进一步研究整合生理和表观遗传输入的机制奠定良好的基础。开发的方法将作为翻译工具，研究与早产相关的围产期应激源和成人疾病的宫内规划。通过执行这项研究计划，候选人将接受所有必要的培训，以成为一名独立的调查员，从事新生儿学的学术研究和教学生涯。至于与公共健康的相关性，现在的大量证据表明，婴儿在出生前发育的环境是规划成人健康的重要因素。例如，在子宫中受到压力的婴儿患心血管疾病和II型糖尿病的风险增加。这一提议试图从分子原因的角度来解释这些神秘的联系。 (摘要结束)

DESCRIPTION (provided by applicant): The Mentored Clinical Scientist Development Award (K08) will allow the candidate to develop into an independent researcher who will conduct both basic and translational research in the field of neonatal medicine. Over the course of this award, Dr. Maltepe will embark upon a didactic and research program designed to provide him with additional training in proteomics and mass spectrometry techniques. He will utilize his mentor's expertise in this area to further explore the epigenetic mechanisms enabling the Hypoxia- Inducible Factor (HIF) family of proteins to regulate hypoxia responses and determine stem cell fate. The specific aims are as follows. (Aim 1) Analyze alterations of well-defined histone modifications in response to hypoxic culture in wild-type and HIF-null mouse embryonic fibroblasts as well as during differentiation in wildtype and HIF-null trophoblast stem cells. (Aim 2) Determine the nature of HIF-interacting chromatin remodeling complexes under the above conditions by utilizing immunoprecipitation, candidate protein immunodetection, and mass spectrometry techniques. (Aim 3) Interrogate the chromatin-associated subproteome for global alterations of histone post-translational modifications under the above conditions using quantitative proteomic and mass spectrometric investigation. (Aim 4) Analyze genome-wide changes in the methylation status of CpG islands under the above conditions using arrays of BAG clones and the methylation-sensitive restriction enzyme Notl. The information obtained from these studies will form a sound foundation to further study the mechanisms responsible for integrating physiological and epigenetic inputs. The developed methodologies will serve as translational tools to study perinatal stressors associated with preterm birth and the intra-uterine programming of adult disease. By carrying out this research plan the candidate will receive all the necessary training to become an independent investigator pursuing an academic research and teaching career in neonatology. As for relevance to public health, a great deal of evidence now suggests that the environment in which a baby develops before birth is an important factor in programming adult health. For example, babies that are stressed in the womb have an increased risk of developing cardiovascular disease and type II diabetes. This proposal seeks to explain these enigmatic associations in term of the molecular causes. (End of Abstract)