CAREER: The Role of F-box-Mediated Protein Degradation in Seed Development

职业：F-box 介导的蛋白质降解在种子发育中的作用

• 批准号: 1750361
• 负责人: Zhihua Hua
• 金额: $ 109.38万
• 依托单位: Ohio University
• 依托单位国家: 美国
• 项目类别: Continuing Grant
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-06-15 至 2024-05-31
• 项目状态: 已结题
• 来源: https://www.nsf.gov/awardsearch/showAward?AWD_ID=1750361&HistoricalAwards=false
• 关键词: CAREER; Role; box; Mediated; Protein

Seeds are important for the propagation of flowering plants and are also the primary source of food for human and animal consumption. Understanding the molecular mechanisms underpinning seed development can help protect plant diversity and secure the ever-increasing demand for food production. Rapid cell division and tissue patterning during the process of seed development result in the production of various short-lived regulatory proteins that are removed in a timely manner. Alterations to these protein removal pathways can cause sterility or a severe reduction in the quantity and quality of seed production. However, currently little is known about the identity of these short-lived proteins and how they are targeted for removal. The goal of this research is to understand the functions of a specific group of proteins, termed the F-box proteins, in regulating protein turnover during seed development. The resulting discoveries can then be translated into crop breeding for developing innovative approaches to manipulate seed production. The interdisciplinary nature of this project will provide a wide array of training opportunties for high school, undergraduate, and graduate students, including those from groups under-represented in science. In addition, this project will develop and promote a "big data" - enabled educational system through classroom teaching, research mentoring in the laboratory, and outreach programs. This training system is critical in modern biological education due to the exponential increase in genomic sequencing. The educational goal of this project is to train diverse students, particularly those from educationally underserved Appalachian areas, to develop computational thinking skills in solving biological questions, thus empowering them to pursue a STEM-based career.The F-box protein family serves as the substrate receptor in Skp1-Cullin 1-F-box (SCF) ubiquitin ligases that target many abnormal and short-lived proteins for ubiquitylation and subsequent degradation by the 26S proteasome. In plants, this family plays a wide range of regulatory roles as suggested by its large expansion in plant genomes and genetic analyses of several members in the family. Unfortunately, little is known about the functions of most F-box proteins. Given the presence of many conserved cellular differentiation programs that coordinate to form a mature seed, the over-arching goal of this project is to uncover the role of evolutionarily-conserved F-box proteins in seed development, using Arabidopsis thaliana as a model. First, RNA-Seq experiments and proteomic analyses will be undertaken to define the global influence of F-box-mediated protein degradation in seed development. Bioinformatic approaches will then be used to construct dynamic regulatory networks between active F-box members, F-box substrates, and transcription factors involved in different seed developmental stages. Second, a novel inducible clustered regularly interspaced short palindromic repeat interference (CRISPRi) approach, combined with conventional T-DNA insertion mutations and over-expression strategies, will be used to uncover the genetic functions of conserved F-box genes in embryogenesis. Third, comprehensive genetic, biochemical, and proteomic approaches will be utilized to fine tune the mechanistic roles of F-box proteins involved in seed development. Fourth, the sequencing data generated from the experiments will be adopted in "big data" - enabled biological education, in which different levels of computational thinking skills will be incorporated to train a diverse group of students. This research is anticipated to contribute in-depth knowledge to both seed biology and the biochemistry of protein ubiquitylation. Innovative materials, methods, and algorithms created in the project will have substantial impacts on furthering the understanding of the role of protein ubiquitylation in gene expression regulation.The Project is jointly funded by Molecular and Cellular Biosciences and Integrative Organismal Systems, with additional support provided by the Directorate's Rule of Life Venture Fund.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.

种子对开花植物的繁殖很重要，也是人类和动物食用的主要食物来源。了解种子发育的分子机制有助于保护植物多样性，确保不断增长的粮食生产需求。在种子发育过程中，快速的细胞分裂和组织模式导致各种短命调节蛋白的产生，这些蛋白被及时去除。这些蛋白质去除途径的改变可能导致不育或种子产量和质量的严重下降。然而，目前对这些短寿命蛋白质的身份以及它们如何被靶向去除知之甚少。本研究的目的是了解一组特定蛋白质的功能，称为F-box蛋白，在调节种子发育过程中的蛋白质周转。由此产生的发现可以转化为作物育种，开发创新方法来操纵种子生产。该项目的跨学科性质将为高中生、本科生和研究生提供广泛的培训机会，包括那些来自科学领域代表性不足的群体的学生。此外，该项目将通过课堂教学、实验室研究指导和外展计划开发和推广“大数据”教育系统。由于基因组测序的指数增长，这种培训系统在现代生物学教育中至关重要。该项目的教育目标是培养多样化的学生，特别是那些来自教育不足的阿巴拉契亚地区的学生，培养他们解决生物学问题的计算思维技能，从而使他们能够从事基于stem的职业。F-box蛋白家族作为Skp1-Cullin 1-F-box （SCF）泛素连接酶的底物受体，该泛素连接酶靶向许多异常和短寿命蛋白，使其泛素化并随后被26S蛋白酶体降解。在植物中，这个家族发挥着广泛的调节作用，这表明它在植物基因组中的大量扩展和家族中几个成员的遗传分析。不幸的是，人们对大多数F-box蛋白的功能知之甚少。鉴于存在许多保守的细胞分化程序，协调形成成熟的种子，本项目的首要目标是揭示进化保守的F-box蛋白在种子发育中的作用，以拟南芥为模型。首先，将进行RNA-Seq实验和蛋白质组学分析，以确定f -box介导的蛋白质降解在种子发育中的全局影响。生物信息学方法将用于构建活跃的F-box成员、F-box底物和参与不同种子发育阶段的转录因子之间的动态调控网络。其次，将采用一种新型的可诱导簇状规则间隔短回文重复干扰（CRISPRi）方法，结合传统的T-DNA插入突变和过表达策略，揭示保守的F-box基因在胚胎发生中的遗传功能。第三，将利用综合的遗传、生化和蛋白质组学方法来微调F-box蛋白在种子发育中的机制作用。第四，从实验中产生的测序数据将被用于“大数据”的生物教育中，在这种教育中，不同水平的计算思维技能将被纳入其中，以培养多样化的学生群体。该研究有望为种子生物学和蛋白质泛素化的生物化学研究提供深入的知识。该项目创造的创新材料、方法和算法将对进一步理解蛋白质泛素化在基因表达调控中的作用产生重大影响。该项目由分子和细胞生物科学和综合生物系统联合资助，并由理事会的生命规则风险基金提供额外支持。该奖项反映了美国国家科学基金会的法定使命，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。

项目成果

To Kill or to Be Killed: How Does the Battle between the UPS and Autophagy Maintain the Intracellular Homeostasis in Eukaryotes?

• DOI: 10.3390/ijms24032221
• 发表时间: 2023-01-22
• 期刊: INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
• 影响因子: 5.6
• 作者: Yu, Peifeng;Hua, Zhihua
• 通讯作者: Hua, Zhihua

Closing target trimming and CTTdocker programs for discovering hidden superfamily loci in genomes

• DOI: 10.1371/journal.pone.0209468
• 发表时间: 2019-07-02
• 期刊: PLOS ONE
• 影响因子: 3.7
• 作者: Hua, Zhihua;Early, Matthew J.
• 通讯作者: Early, Matthew J.

Adaptive and degenerative evolution of the S-Phase Kinase-Associated Protein 1-Like family in Arabidopsis thaliana

• DOI: 10.7717/peerj.6740
• 发表时间: 2019-04-12
• 期刊: PEERJ
• 影响因子: 2.7
• 作者: Hua, Zhihua;Gao, Zhenyu
• 通讯作者: Gao, Zhenyu

Diversifying Evolution of the Ubiquitin-26S Proteasome System in Brassicaceae and Poaceae

• DOI: 10.3390/ijms20133226
• 发表时间: 2019-07-01
• 期刊: INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
• 影响因子: 5.6
• 作者: Hua, Zhihua;Yu, Peifeng
• 通讯作者: Yu, Peifeng