The Unfolded Protein Response as an Organizer of Chemosensory Response

描述（由申请人提供）：嗅觉受体（OR）的选择，数千个可用的哺乳动物OR等位基因中的一个的转录激活是一个知之甚少的过程。我们之前证明，在响应OR翻译时，er驻留激酶Perk使翻译起始因子eif2a磷酸化，引发最终稳定OR选择的信号。遗传实验表明，这种反馈信号依赖于转录因子ATF5核异构体翻译的短暂但普遍的衰减和特异性的翻译上调。核ATF5的产生增强腺苷酸环化酶3 （Adcy3）的转录，从而缓解OR诱导的内质网应激，抑制组蛋白去甲基化酶LSD1的表达，使嗅觉神经元终末分化，并使所选择的OR永久表达。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，试图提供这些问题的答案，并为这种未折叠蛋白质反应途径的新用途提供机制见解。具体来说，我们提出了生化和遗传实验，旨在揭示OR蛋白是否直接与Perk相互作用，并绘制出负责Perk激活的确切肽。此外，我们的目标是阐明ATF5作为转录调控因子的作用机制，并探索两种不同的核异构体的功能，根据我们的初步遗传分析，这两种核异构体在调节或选择和嗅觉神经元分化中发挥不同的作用。最后，我们提出了旨在剖析该信号通路的最后一步的实验，即内质网应激的缓解和or诱导的Perk信号的终止。及时终止未折叠蛋白反应的这一臂对于OR表达的稳定与该途径的启动一样重要，我们假设OR特异性伴侣蛋白在阻止ER中OR- perk相互作用中起着关键作用。遗传和生化实验将揭示这些蛋白质的身份，并检查它们在or引发的反馈中的作用。我们的实验将为发现最大的哺乳动物基因家族以来一直保持神秘的过程提供新的见解，并将揭示可能适用于许多其他化学受体家族介导与外界沟通的调节原理。

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.