NSF-BSF: Control of molecular, cellular, and organismal phenotypes by the transcription factor NFAT5

这个项目调查鱼类如何应对它们生活的水中盐分施加的压力。理解这一影响生存、生长和繁殖的重要过程的关键是研究鱼类如何感知环境盐度，以及它们如何产生内部信号，协调调节分子网络以抵消压力。该项目使用新的分子、遗传和生化工具，结合定制的计算模型，检查涉及的分子及其相互作用。具体地说，通过研究对盐胁迫的反应，该项目有可能帮助从总体上了解不同生物组织水平对环境胁迫的反应。通过研究罗非鱼的这些过程，罗非鱼是全球第二大水产养殖物种，这项工作将对基本的食物供应系统产生积极影响。此外，罗非鱼还可以作为水生脊椎动物应激生物学的模式系统。该项目生成工具和知识，通过相关的公共储存库和交流平台与更广泛的科学界共享。该项目包括对所有职业阶段的研究人员进行详细的跨学科培训，重点是在尖端实验和网络建模方法方面未被充分代表的少数群体。该项目确定了与罗非鱼耐盐性至关重要的分子、细胞和生物表型相关的调控网络。研究假设是转录因子NFAT5是控制这些网络的中央协调器。将测试NFAT5与盐胁迫(高渗透压)期间的适应性表型(耐受性、生长、繁殖力)之间可能存在的因果联系。该项目基于先前的研究，阐明了分子机制和生成工具，用于研究鱼类适应性表型的机制原因。NFAT5的功能将通过研究依赖于NFAT5的分子网络以及细胞和有机体的表型，在系统规模的整体水平上确定。重点是将高渗的、依赖于NFAT5的转录组和蛋白质组网络的调节与生长、生存、增殖和繁殖联系起来。将确定转录组和蛋白质组调节耦合程度的潜在规则，并在不同的细胞/组织类型和基因组背景下测试它们的普遍性。细胞系将被用于高通量基因打靶和细胞表型鉴定，以确定产生NFAT5+和NFAT5转基因鱼类用于生物表型鉴定的最佳靶标。更广泛的影响包括促进预测和减轻盐度胁迫对具有高商业价值的生物的后果。用于应对压力的一般适应性策略将被揭示并广泛分享。工具(工程细胞系、质粒库、蛋白质组学文库)和产生的想法将通过GitHub等交流平台和在会议上分享。将为研究生、REU学生、博士后、萨克拉门托地区社区大学学生、K-12学生和教师以及公众提供跨学科培训和推广。这一美国/以色列项目由美国国家科学基金会和以色列双国科学基金会支持。该奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

This project investigates how fish cope with the stress imposed by salt in the water in which they live. Key to understanding this important process -- which impacts survival, growth, and reproduction -- is to examine how the fish sense environmental salinity and how they generate, in response, internal signals that orchestrate the regulation of molecular networks that counteract the stress. The project examines the molecules involved, and their interactions, using novel molecular, genetic, and biochemical tools in combination with custom-built computational models. By studying responses to salt stress, specifically, this project has the potential to help understand, in general, responses to environmental stress at different levels of biological organization. By examining these processes in tilapia, the second-most important aquaculture species globally, this work will positively impact an essential food supply system. In addition, tilapia can serve as a model system for stress biology in aquatic vertebrate species. This project generates tools and knowledge shared with the broader scientific community via pertinent public repositories and communication platforms. It includes elaborate cross-disciplinary training of researchers at all career stages, with emphasis on underrepresented minorities, in cutting-edge experimental and network modeling approaches.This project identifies regulatory networks that are causally linked to molecular, cellular, and organismal phenotypes critical for salinity tolerance in tilapia, Oreochromis mossambicus. The research hypothesis is that NFAT5, a transcription factor, is a central orchestrator for controlling these networks. Putative causal links between NFAT5 and adaptive phenotypes (tolerance, growth, fecundity) during salinity stress (hyperosmolality) will be tested. The project is based on prior research elucidating molecular mechanisms and generating tools for studying mechanistic causes of adaptive phenotypes in fish. NFAT5 functions will be determined on a systems-scale, holistic level by investigating molecular networks and cellular and organismal phenotypes that depend on NFAT5. The emphasis is on linking the hyperosmotic, NFAT5-dependent regulation of transcriptome and proteome networks with growth, survival, proliferation, and reproduction. Rules underlying the extent of coupling of transcriptome versus proteome regulation will be determined and their universality tested in different cell-/tissue types and genomic backgrounds. Cell lines will be utilized for high-throughput gene targeting and cellular phenotyping to identify the best targets for generating NFAT5+ and NFAT5- transgenic fish for organismal phenotyping. Broader impacts include the facilitation of prediction and mitigation of consequences of salinity stress on organisms of high commercial value. General adaptive strategies used to cope with stress will be revealed and shared broadly. Tools (engineered cell lines, plasmids, proteomics libraries) and ideas generated will be shared via communication platforms such as GitHub and at conferences. Cross-disciplinary training and outreach will be provided to graduate students, REU students, a postdoc, Sacramento area community college students, K-12 students and teachers, and the public.This US/Israel project is supported by the US National Science Foundation and the Israeli Binational Science Foundation.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.