CAREER: Arabidopsis Argonaute10-protein Interactome

智力优势：Argonaute （AGO）蛋白是rna介导的基因沉默的关键效应物，通过与mirna结合在转录或转录后水平抑制靶基因的表达。虽然以前认为AGOs只抑制miRNA靶基因，但PI的研究小组最近发现拟南芥AGO10特异性地隔离了单一miRNA类miR166，以积极调节其靶标。这种调控机制尚不清楚，但可能涉及与AGO10相互作用的蛋白。初步研究发现了几个新的AGO10辅因子，包括两个丝氨酸和精氨酸富集剪接因子（srsf）和一个AGO10相互作用的外核糖核酸酶（ager1）等。这些新发现将RNA沉默与许多以前未被发现的生物学事件（如mRNA剪接和ago通道的miRNA衰变）联系起来。为了研究这些新的AGO10调控模式，我们将追求两个特定的目标：1)研究AGO10与SRSFs之间的生化相互作用及其生物学后果；2)研究ager1对AGO10-miR166功能的影响。研究结果将揭示AGO10/miR166功能是如何被调节的，并通过miRNA功能揭示基因调控网络的新复杂性。更广泛的影响：该研究为本科生和研究生在广泛的领域提供了独特和先进的培训机会。研究和教育目标通过两种策略整合到该CAREER提案中：1)招收和培训生物化学和遗传领域的女性和少数民族本科生；2)在课堂和研究环境中现代化和提高RNA生物学研究生培训的质量。这些努力将有助于提供具有创造力和高技能的劳动力，以支持美国经济。

Intellectual Merit: Argonaute (AGO) proteins are key effectors of RNA-mediated gene silencing and act by binding to miRNAs to repress the expression of target genes at transcriptional or posttranscriptional levels. Whereas AGOs were previously thought to exclusively repress miRNA target genes, the PI's group recently discovered that Arabidopsis AGO10 specifically sequesters a single miRNA class, miR166 to positively regulate their targets. The mechanism of this regulation is unclear, but it may involve proteins that interact with AGO10. Preliminary studies identified several novel cofactors of AGO10, including two serine and arginine enriched splicing factors(SRSFs) and an AGO10-Interacting exoribonuclease (AGIER1), among others. These new findings interconnect RNA silencing with many previously unappreciated biological events such as mRNA splicing and AGO-channeled miRNA decay. To investigate these novel modes of AGO10 regulation, two specific objectives will be pursued: 1) Investigate the biochemical interaction between AGO10 and the SRSFs and its biological consequences; 2) Investigate how AGIER1 affects AGO10-miR166 function. Results will establish how AGO10/miR166 function is modulated and will reveal new layers of complexity in gene regulatory networks through miRNA function. Broader Impacts: The research provides unique and advanced training opportunities for undergraduates, and graduate students in broad areas. Research and educational objectives are integrated in this CAREER proposal through two strategies: 1) to recruit and train female and minority undergraduates for graduate studies in biochemical and genetic areas; and 2) to modernize and improve the quality of graduate training in RNA biology in classroom and research settings. These efforts will contribute to providing a creative and highly skilled workforce to support the U.S. economy.