Developmental Requirements of SWI/SNF Chromatin Remodeling Complexes in Porcine Embryos

 描述(申请人提供)：美国估计有600万人患有不孕症，约占育龄人口的10%。治疗人类不孕不育可以涉及辅助生殖技术，通常包括体外受精和胚胎培养。众所周知，与在体内生产的胚胎相比，体外生产的哺乳动物胚胎具有更低的发育能力。我们还知道，在实验室培养的胚胎可以表现出表观遗传变化(DNA甲基化和共价组蛋白修饰)、基因表达异常和胎儿生长模式受扰。尽管已知体外胚胎操作与扰动的表观遗传和转录状态之间的联系，但人们对卵裂阶段胚胎的表观遗传变化的调节机制以及这些表观遗传变化如何导致发育失败知之甚少。为了达到改善体外操作人类胚胎结局的长期目标，该建议涉及两个具体的目标：1)确定SWI/SNF染色质重塑亚基进入猪卵母细胞核和卵裂期胚胎的卵裂球的机制；2)确定卵裂期胚胎中离散的SWI/SNF染色质重塑复合体亚基的发育需求。这些目标将通过一系列实验来实现，这些实验涉及去化和过度表达方法，以确定SWI/SNF染色质重塑酶的调节成分是如何运输到卵裂期胚胎的细胞核的。猪胚胎由于在胚胎发育的早期阶段与人类胚胎在生理上的相似性而被用作模型系统。这些研究的发现将对人类生殖医学产生影响，并将导致改进的胚胎操作和培养方法的开发。

 DESCRIPTION (provided by applicant): Infertility affects an estimated 6 million people in the United States, roughly 10% of the population of reproductive age. Treating human infertility can involve assisted reproductive technologies, which often include in vitro fertilization and embryo culture. It is well-established that mammalian embryos produced in vitro possess reduced developmental competence as compared to embryos produced in vivo. It is also known that embryos cultured in the laboratory can display epigenetic changes (both in DNA methylation and covalent histone modifications), aberrant gene expression and perturbed patterns of fetal growth. Despite the known links between in vitro embryo manipulation and perturbed epigenetic and transcriptional states, very little is known about the mechanisms by which epigenetic changes are mediated in cleavage stage embryos and how these epigenetic changes lead to developmental failure. To move toward the long term goal of improving the outcome for human embryos manipulated in vitro, two specific aims are addressed in this proposal; the aims are as follows: 1) Determine the mechanism by which SWI/SNF chromatin remodeling subunits access the nuclei of porcine oocytes and blastomeres of cleavage stage embryos, and 2) determine the developmental requirements of discrete SWI/SNF chromatin remodeling complex subunits in cleavage stage embryos. These aims will be addressed through a series of experiments that involve abalation and overexpression approaches to determine how regulatory components of the SWI/SNF chromatin remodeling enzymes are trafficked to the nuclei in cleavage stage embryos. The porcine embryo is used as the model systems because of its physiologic similarity to the human embryo during the early stages of embryonic development. The findings from these studies will have an impact on human reproductive medicine and will lead to development of improved methods for manipulating and culturing embryos.