Nuclear action of a novel fish kisspeptin receptor isoform

该项目解决了蛋白质在基因复制事件后发挥额外作用的有趣话题。它专注于kisspeptin激素及其受体，这是一种在生殖控制中起关键作用的激素信号通路。Kisspeptin受体是G蛋白偶联受体（GPCR）家族的成员，该家族是最大的蛋白质家族。GPCR信号的教科书观点是，细胞表面的激素结合引起跨膜受体蛋白的构象变化，并促进其与G蛋白的相互作用，以改变细胞内或细胞膜中的酶和离子通道的活性，从而导致特异性细胞反应。越来越多的证据表明，这种观点可能过于简单化，因为已经发现许多GPCR不仅位于细胞膜上，而且位于细胞内的各个隔室中。研究人员发现，斑马鱼kisspeptin受体B基因编码几种蛋白质，其中一种位于细胞核内，具有核活性。这些发现提出了一个令人兴奋的可能性，以前未被承认的核功能的GPCR。本项目旨在验证这一假设。这些结果将为理解kisspeptin途径如何调节鱼类繁殖提供新的见解，并应导致在水产养殖业中的应用。该项目将为不同层次的学生提供培训机会，并将通过让K-12学生和公众参与教育推广活动来造福社会。kisspeptin配体（KISS 1）和受体（KISS 1 R）激素途径在哺乳动物生殖的神经内分泌控制中起着关键作用。哺乳动物KISS 1-KISSR通路也具有非生殖功能，但机制知之甚少。最近的研究表明，斑马鱼有两个kisspeptin受体基因。虽然斑马鱼kiss 1 ra基因编码一个单一的G蛋白偶联受体（GPCR）在大脑中表达，kiss 1 rb基因编码全长GPCR以及四个选择性剪接亚型。这些截短的同种型，称为kiss 1 rb衍生蛋白（KRBDP），在大脑和许多外周组织中表达。功能分析结果表明，这些KRBDP转录本可以翻译成蛋白质产物，但没有位于质膜上。有趣的是，KRBDP 3在细胞核中被检测到，并且在体外测试时具有配体非依赖性的反式激活活性。该项目将测试KRBDP 3定位于细胞核并调节脑和/或外周组织中的基因表达的假设。这些结果将阐明一种新的GPCR亚型在细胞核中的功能，并确定其靶基因。该项目还将在鱼类模型中揭示重复的Kiss 1-Kiss 1 r通路在大脑和外周组织中的体内功能。该项目包含技术创新，将使剖析不同选择性剪接基因产物的功能并可视化其体内表达和亚细胞定位成为可能。它将为不同层次的学生提供高质量的研究培训，并与密歇根大学自然历史博物馆合作开展公共教育活动。该奖项反映了NSF的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

The project addresses the interesting topic of proteins that take on additional roles after a gene duplication event. It focuses on the kisspeptin hormone and its receptor, a hormonal signaling pathway that plays key roles in the control of reproduction. The kisspeptin receptor is a member of the G-protein coupled receptor (GPCR) family, the largest protein family. The textbook view of GPCR signaling is that hormone binding at the cell surface causes a conformational change in the transmembrane receptor protein and promotes its interaction with G proteins to alter the activity of enzymes and ion channels inside the cell or in the cell membrane, leading to specific cellular responses. There is increasing evidence that this view may be too simplistic, because a number of GPCRs have been found to be located not only on the cell membrane but also in various compartments within cells. The researchers have discovered that the zebrafish kisspeptin receptor b gene encodes several proteins, one of which is located in the nucleus and has nuclear activity. These findings present the exciting possibility of a previously unrecognized nuclear function of a GPCR. This project is designed to test this hypothesis. The results will provide new insights into the understanding of how the kisspeptin pathway regulates fish reproduction, and should lead to applications in the aquaculture industry. This project will provide training opportunities for a diverse group of students at all levels, and will benefit society by engaging K-12 students and the general public in educational outreach activities.The kisspeptin ligand (KISS1) and receptor (KISS1R) hormonal pathway plays key roles in the neuroendocrine control of reproduction in mammals. The mammalian KISS1-KISSR pathway also has non-reproductive functions with poorly understood mechanisms. Recent studies suggest that the zebrafish has two kisspeptin receptor genes. While the zebrafish kiss1ra gene encodes a single G-protein coupled receptor (GPCR) expressed in the brain, the kiss1rb gene encodes a full-length GPCR as well as four alternatively spliced isoforms. These truncated isoforms, termed kiss1rb-derived proteins (KRBDPs), are expressed in the brain and in many peripheral tissues. Functional analysis results suggest that these KRBDP transcripts can be translated into protein products, but none are located on the plasma membrane. Intriguingly, KRBDP3 is detected in the nucleus and possesses ligand-independent transactivation activity when tested in vitro. This project will test the hypothesis that KRBDP3 is localized in the nucleus and regulates gene expression in the brain and/or peripheral tissues. The results will elucidate the functions of a novel GPCR isoform in the nucleus and identify its target genes. This project will also uncover the in vivo functions of the duplicated Kiss1-Kiss1r pathway in the brain and peripheral tissues in a fish model. This project contains technical innovations that will make it possible to dissect the functions of different alternatively spliced gene products and to visualize their expression and subcellular localization in vivo. It will provide high-quality research training for a diverse group of students at all levels, and public educational activities in partnership with the University of Michigan Museum of Natural History.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.