RNA editing-mediated modulation of serotonin 2C receptor protein expression

描述（由申请人提供）：编码5-羟色胺（5 HT 2C）受体2C亚型的转录物可以通过多达5个腺苷至肌苷RNA编辑事件进行修饰，产生多达24种蛋白质同种型，这些蛋白质同种型在其受体：G-蛋白偶联功效和组成型活性方面不同。5 HT 2C mRNA的编辑变异已在情感障碍中被鉴定，包括焦虑症、精神分裂症和与自杀相关的抑郁症。在最近的工作中，我们证明了仅表达完全编辑的5 HT 2C受体亚型的转基因小鼠显示出与Prader-Willi综合征一致的表型变化，Prader-Willi综合征是一种人类发育障碍，其特征在于最初无法茁壮成长，认知缺陷，躯体生长减少，新生儿肌肉张力减退和与病态肥胖相关的摄食过多。有趣的是，与野生型动物相比，这些突变小鼠的5 HT 2C受体密度增加了40- 70倍，但稳态mRNA水平保持不变。因此，RNA编辑通过未表征的转录后机制对5 HT 2C蛋白的表达具有显著影响。本提案的目的是研究基于RNA编辑的5 HT 2C受体密度改变的分子机制。在第一个目标中，将使用体内和体外模型系统来确定5 HT 2C编辑对受体稳定性的影响。还将检查编辑对5 HT 2C运输和/或配体活化以及随后的受体稳定性的影响。在目标2中，将研究未编辑和完全编辑的5 HT 2C亚型之间的翻译的潜在差异。此外，MBII-52的翻译效果，一个小的核仁RNA（snoRNA），可以影响5 HT 2C选择性剪接，将在编辑和非编辑的5 HT 2C mRNA进行研究。目的3表示在整个小鼠脑中相对5 HT 2C蛋白同种型水平的定量分析。预计这些研究将为与5 HT 2C功能改变相关的人类疾病提供新的见解，其中关于编辑的受体亚型表达的推断仅基于mRNA的分析。 公共卫生相关性：这项研究计划将增加我们对哺乳动物神经系统中5-羟色胺2C受体（5 HT 2C）功能多样性和调节回路的了解。我们的新发现有可能重新定义5 HT 2C相关疾病的研究，诊断和治疗，包括精神分裂症，抑郁症，焦虑和成瘾。

DESCRIPTION (provided by applicant): Transcripts encoding the 2C-subtype of serotonin (5HT2C) receptor can be modified by up to five adenosine-to- inosine RNA editing events, generating as many as 24 protein isoforms that differ in their receptor:G-protein coupling efficacy and constitutive activity. Variations in the editing of 5HT2C mRNAs have been identified in affective disorders including anxiety, schizophrenia and depression associated with suicide. In recent work, we demonstrate that genetically-modified mice solely expressing the fully edited isoform of the 5HT2C receptor display phenotypic changes consistent with Prader-Willi Syndrome, a human developmental disorder characterized by an initial failure to thrive, cognitive deficits, decreased somatic growth, neonatal muscular hypotonia, and hyperphagia associated with morbid obesity. Interestingly, these mutant mice exhibit an unprecedented 40- to 70-fold increase in 5HT2C receptor density compared to wild-type animals, yet the steady-state mRNA levels remain unchanged. Thus, RNA editing has dramatic consequences on the expression of 5HT2C protein through uncharacterized post-transcriptional mechanism(s). The objectives of this proposal are to investigate the molecular mechanism(s) underlying RNA editing-based alterations in 5HT2C receptor density. In the first aim, the effects of 5HT2C editing on receptor stability will be determined using both in vivo and in vitro model systems. The effects of editing on 5HT2C trafficking and/or ligand activation and subsequent stability of the receptor also will be examined. In aim 2, potential differences in translation among non-edited and fully edited 5HT2C isoforms will be investigated. Furthermore, translation effects of MBII-52, a small nucleolar RNA (snoRNA) that can affect 5HT2C alternative splicing, will be investigated on both edited and non-edited 5HT2C mRNAs. Aim 3 represents a quantitative analysis of relative 5HT2C protein isoform levels in whole mouse brain. It is anticipated that these studies will provide novel insights into human disorders related to altered 5HT2C function, where inferences about the expression of edited receptor isoforms have been based solely upon analyses of mRNA. PUBLIC HEALTH RELEVANCE: This research plan will increase our understanding of the functional diversity and regulatory circuits of the serotonin 2C receptor (5HT2C) in the mammalian nervous system. Our novel findings have the potential to redefine research, diagnosis and treatment of 5HT2C-related disorders including schizophrenia, depression, anxiety and addiction.