RNA Binding Proteins in Complex Neurological Disease

 描述(申请人提供)：许多因素使基因复杂的疾病变得复杂。传统上，它们被定义为多种遗传变异和非遗传因素之间的相互作用。基因组测序和种内变异方面的进展引起了人们对从人类和模式生物中识别多基因变异的极大兴趣，并取得了一些成功。但人们不能忽视生理复杂性的重要性；当在复杂的生物系统中运作时，即使是孟德尔式的变体也可能造成严重破坏。对于功能性表型，例如中枢神经系统的兴奋性障碍，这一概念还没有得到充分的研究。当然，癫痫在基因上是复杂的，但作为大脑的典型兴奋性障碍，它也是研究其他更难破解的功能性障碍的典范，如自闭症和精神分裂症，这些疾病的核心也可能是兴奋病理。神经元的兴奋性主要由离子通道和转运体、神经递质受体和突触蛋白等分子决定，它们控制膜电位和突触信号，以实现兴奋和抑制的适当平衡。虽然编码这些分子的基因中的顺式变异可以导致特定的表型，但调节它们表达的反式因子必须是维持这种平衡在更高、协调水平上的关键。我们以前鉴定并鉴定了Celf4(以前称为Brunol4)的亚型和零基因类型，它编码了Bruno/CUGBP/CELF RNA结合蛋白家族中的一个脑特异性成员。Celf4突变体有复杂的癫痫发作障碍和其他神经表型，如多动、轻度肥胖和异常社交。最近，人类CELF4缺乏症揭示了这些症状和其他症状，如智力残疾。在我们最初的资助期间，我们发现CELF4与非常高密度的RNA颗粒联系最紧密，并以兴奋性神经元中的大量mRNAs为靶标。突触功能涉及许多靶点，它们往往在突变小鼠的神经元内处于失调状态--在各个方向上，但有一种在细胞体以外表达增加的趋势。这些发现与CELF4在控制“翻译沉默”中的作用是一致的，可能是在局部、亚细胞水平。我们还获得了CELF4通过增加钠通道Nav1.6的表达而对内源性神经元过度兴奋起作用的证据，以及可能由于突触蛋白调节失调而导致的系统范围内平衡可塑性受损的证据。我们的假设是，这些影响的组合导致了全面的疾病。在接下来的5年里，我们的研究将解决两个共同努力解决这一想法的流行主题。第一个研究讨论了目标mRNA3‘-UTR中CELF4结合基序的模式，以及改变基序对mRNA丰度、定位和翻译的影响，无论是在转录组范围内，还是在选定的靶点上。第二个主题使用体内突变来评估个体靶点对复杂神经表型的多基因病因学的贡献。

 DESCRIPTION (provided by applicant): Many factors make genetically complex diseases complex. Classically they are defined as an interaction between multiple genetic variants and non-genetic factors. Progress in genome sequencing and within-species variation has generated much interest in identifying polygenic variants from human and model organisms, with some success. But one cannot lose sight of the importance of physiological complexity; even Mendelian variants can wreak havoc when operating in a complex biological system. For functional phenotypes, such as excitability disorders of the CNS, this concept is understudied. Epilepsy is genetically complex to be sure, but as the canonical excitability disorder of the brain it also serves as a leading example for approaching other, harder-to-crack functional disorders, such as autism and schizophrenia, that are also likely to have excito-pathology at their cores. Neuronal excitability is determined primarily by molecules, such as ion channels and transporters, neurotransmitter receptors, and synaptic proteins, controlling membrane potential and synaptic signaling in order to achieve an appropriate balance of excitation and inhibition. Although cis-variants in genes encoding these molecules can lead to specific phenotypes, trans-factors that regulate their expression must be critical for maintaining this balance at a higher, coordinated level. We previously identified and characterized hypomorphic and null genotypes in Celf4 (formerly known as Brunol4), encoding a brain-specific member of the BRUNO/CUGBP/CELF family of RNA binding proteins. Celf4 mutants have a complex seizure disorder and other neurological phenotypes, such as hyperactivity, mild obesity and abnormal social interaction. Very recently human CELF4 deficiency revealed these and additional symptoms, such as intellectual disability. In our initial funding period, we found that CELF4 is most tightly associated with very high-density RNA granule particles and targets a vast number of mRNAs in excitatory neurons. Many targets are involved in synaptic functions, and they tend to be dysregulated within neurons of mutant mice - in all directions, but with a tendency towards increased expression away from the cell body. These findings are consistent with a role for CELF4 in control "translational silencing" perhaps at local, subcellular levels. We also obtained evidence for CELF4 effects on intrinsic neuronal hyperexcitation, via increased expression of sodium channel Nav1.6, and system-wide dysregulation via impaired homeostatic plasticity presumably due to dysregulation of synaptic proteins. Our hypothesis is that the combination of such effects accounts for full-blown disease. In the next 5 years, our research will address two prevailing themes that work together to address this idea. The first addresses the pattern of CELF4 binding motifs within the 3'-UTR of target mRNAs, and the consequences of altering the motifs on mRNA abundance, localization and translation, both transcriptome-wide and for selected targets. The second theme uses in vivo mutagenesis to assess the contribution of individual targets to the multigenic etiology of complex neurological phenotypes.