Understanding the Role of Nuclear Architecture in Olfactory Receptor Choice

描述（由申请人提供）：嗅觉受体（OR）选择，即数千种可用哺乳动物OR等位基因之一的转录激活，是一个知之甚少的过程。我们先前证明了OR基因在少数OR特异性异染色质病灶中的核内和染色体间聚集有助于有效的OR沉默，保留了OR表达的单基因和单等位基因性质。在这里，我们研究的假设，复杂的染色体间协会也负责一个单一的OR等位基因的转录激活。使用高通量表观遗传和遗传方法，我们确定了一组新的OR增强子，支持报告基因在斑马鱼和小鼠的嗅觉感觉神经元的重要部分表达。我们的初步数据表明，这些增强子可能协同激活OR表达，在嗅觉感觉神经元的核中产生结构和功能奇点。我们提出了一系列的实验，将映射和量化的染色体间协会的新确定的增强子使用成像和Hi-C的方法。此外，我们提出了遗传损失和获得的功能实验，将测试是否或增强剂协同作用对一个单一的OR等位基因的激活。最后，我们试图研究OR增强子在其功能和核组织中的不寻常表观遗传特征的贡献。我们的实验将提供新的见解核结构在基因表达中的作用，并将揭示参与体内细胞多样性的产生的分子机制。

DESCRIPTION (provided by applicant): Olfactory receptor (OR) choice, the transcriptional activation of one out of thousands of available mammalian OR alleles, is a poorly understood process. We previously demonstrated that the nuclear intra- and inter- chromosomal aggregation of OR genes in a few, OR-specific, heterochromatic foci contributes to the efficient OR silencing, preserving the monogenic and monoallelic nature of OR expression. Here, we examine the hypothesis that complex interchromosomal associations are responsible also for the transcriptional activation of a single OR allele. Using high throughput epigenetic and genetic approaches we identified a set of novel OR enhancers that support reporter expression in significant fractions of olfactory sensory neurons in zebrafish and mice. Our preliminary data suggest that these enhancers might work in concert for the activation of OR expression, creating a structural and functional singularity in the nuclei of olfactory sensory neurons. We propose a series of experiments that will map and quantify the interchromosomal associations of the newly identified enhancers using imaging and Hi-C approaches. Moreover, we propose genetic loss-, and gain-of- function experiments that will test whether OR enhancers act synergistically towards the activation of a single OR allele. Finally, we seek to investigate the contribution of the unusual epigenetic signature of OR enhancers in their function and nuclear organization. Our experiments will provide novel insight into the role of nuclear architecture in gene expression and will uncover molecular mechanisms involved in the generation of cellular diversity in vivo.