The Unfolded Protein Response as an Organizer of Chemosensory Response

描述(申请人提供)：嗅觉受体(OR)选择，数以千计的哺乳动物OR等位基因中的一个转录激活是一个鲜为人知的过程。我们先前证明，在对OR翻译的响应中，内质网驻留的激酶Perk磷酸化翻译起始因子eif2a，引发一个最终稳定OR选择的信号。遗传学实验表明，这种反馈信号依赖于转录因子ATF5的核异构体翻译的瞬时但普遍的减弱和特异性的上调。核ATF5的产生促进了腺酰环化酶3(Adenylyl Cyclase 3，Adcy3)的转录，从而缓解了OR诱导的内质网应激，抑制了组蛋白去甲基酶LSD1的表达，从而允许嗅觉神经元的终末分化，使所选择的OR的表达永久化。这些观察结果对该信号通路的分子原理提出了重要的问题。在这里，我们提出了一些实验，试图为这些问题提供答案，并为这种未折叠蛋白质反应途径的新用途提供机制上的洞察。具体地说，我们提出了生化和遗传学实验，旨在揭示OR蛋白是否直接与PERK相互作用，并绘制负责PERK激活的确切多肽。此外，我们的目的是阐明ATF5作为转录调节因子的作用机制，并根据我们的初步遗传分析，探索两种不同的核异构体在调节OR选择和嗅觉神经元分化中的不同作用。最后，我们提出了实验，旨在剖析这一信号通路的结束步骤，即缓解内质网应激和终止OR诱导的PERK信号。及时终止未折叠蛋白反应的这一臂对于OR表达的稳定与这一途径的启动同样关键，我们假设OR特异性伴侣在阻止ER中的OR-Perk相互作用中发挥关键作用。遗传和生化实验将揭示这些蛋白质的特性，并检查它们在OR引发的反馈中的作用。我们的实验将为这一过程提供新的见解，这一过程自发现最大的哺乳动物基因家族以来一直是个谜，并将揭示可能适用于许多其他化学感受器家族的调节原理，这些家族调节与外部世界的沟通。

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 in response to OR translation, the ER-resident kinase Perk phosphorylates the translation initiation factor eif2a, eliciting a signal that culminates in the stabilization of OR choice. Genetic experiments suggest that this feedback signal depends upon the transient but general attenuation of translation and the specific upregulation of translation of the nuclear isoform of transcription factor ATF5. Production of nuclear ATF5 enhances the transcription of Adenylyl cyclase 3 (Adcy3), which relieves the OR-induced ER stress and represses the expression of histone demethylase LSD1, allowing the terminal differentiation of olfactory neurons and making the expression of the chosen OR permanent. These observations pose significant questions regarding the molecular principles of this signaling pathway. Here, we propose experiments that seek to provide answers to these questions and to offer mechanistic insight into this novel use of the unfolded protein response pathway. Specifically, we propose biochemical and genetic experiments aiming to reveal whether OR proteins interact directly with Perk and to map the exact peptides responsible for Perk activation. Furthermore, we aim to elucidate the mechanism of action of ATF5 as transcriptional regulator and to explore the function of two distinct nuclear isoforms that, according to our preliminary genetic analysis, play different roles in the regulation of OR choice and the differentiation of olfactory neurons. Finally, we propose experiments that aim to dissect the concluding step of this signaling pathway, which is the relief of ER stress and the termination of OR-induced Perk signaling. Timely termination of this arm of the unfolded protein response is as critical for the stabilization of OR expression as the initiation of this pathway an we hypothesize that OR-specific chaperones play a critical role in preventing OR-Perk interactions in the ER. Genetic and biochemical experiments will reveal the identity of these proteins and examine their role in the OR-elicited feedback. Our experiments will provide novel insight into a process that has remained enigmatic since the discovery the largest mammalian gene family and will reveal regulatory principles that likely apply to many other chemoreceptor families mediating communication with the outside world.