Research Initiation Award:Transcriptome Analysis of Microtubule Actin Crosslinking Factor 1 (MACF1) Stage-Specific oocytes in Zebrafish

通过研究启动奖的历史黑人学院和大学本科课程(HBCU-UP)为历史黑人学院和大学的初级和职业生涯中期教师提供支持，这些教师正在建立新的研究项目或重新定向和重建现有的研究项目。预计该奖项有助于进一步提高教师的研究能力和有效性，并改善所在机构的研究和教学。授予科平州立大学的这一奖项为建立斑马鱼实验室提供了机会，该实验室研究卵子发生的机制。具体地说，该项目旨在研究微管肌动蛋白交联因子1(MACF1)在Balbiani小体(BB)分解中的功能。该奖项还包括培养研究生、本科生和高中生，从而为下一代科学家做出贡献，他们可以为科学创新、创造力和生产力做出贡献。胚轴首先在卵子发生期间由卵母细胞中建立的极性线索确定，然后再激活胚胎基因组。这种明显的不对称性是由一种植物局部结构决定的，称为巴尔比亚尼小体(BB)。BB在从脊椎动物到无脊椎动物的卵母细胞中都是保守的，由细胞器和蛋白质组成的聚集体，包括RNA结合蛋白。然而，BB在卵子发生和解体过程中建立极性的机制尚不清楚。有证据表明，麦哲伦(Magellan，MGN)突变导致卵极性缺陷，其特征是BB增大和细胞核错位。MGN突变体特别是破坏了微管肌动蛋白交联因子(Macf1a)基因，该基因在其他极化细胞中起作用。这项拟议研究的目标将集中在卵子发生的早期阶段，这些阶段会产生成熟和可存活的卵母细胞，以便成功受精。为了解决Bb的拆解及其与MACF1a的相互作用，本研究将1)利用下一代测序(NGS)技术来测量MACF1a阶段特定卵母细胞中转录产物的差异表达；2)在超微结构水平上显示MACF1a蛋白与Bb的结合。拟议的项目将首次发现斑马鱼母性效应突变体(Macf1)的转录组，这可能会导致更深入地了解控制极性的基因，同时提供MACF1蛋白/Balbiani身体的高分辨率特征。这一奖项反映了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。

The Historically Black Colleges and Universities Undergraduate Program (HBCU-UP) through Research Initiation Awards provide support for junior and mid-career faculty at Historically Black Colleges and Universities who are building new research programs or redirecting and rebuilding existing research programs. It is expected that the award helps to further the faculty member's research capability and effectiveness and improve research and teaching at the home institution. This award to Coppin State University provides an opportunity to establish a zebrafish laboratory that investigates the mechanisms underlying oogenesis. Specifically, the project aims to examine the function of Microtubule Actin Crosslinking Factor 1 (MACF1) in Balbiani body (Bb) disassembly. With this award, efforts also include the training of graduate, undergraduate, and high school students, thereby contributing to the next generation of scientists who can contribute to scientific innovation, creativity and productivity.The embryonic axis is first determined during oogenesis by polarity cues established in the oocyte, preceding embryonic genome activation. This distinct asymmetry is determined from a vegetally localized structure, called the Balbiani body (Bb). The Bb is conserved in oocytes from vertebrates to invertebrates and is composed of an aggregation of organelles and proteins including RNA-binding proteins. However, the mechanism by which the Bb establishes polarity during oogenesis and disassembles is unknown. There is evidence to demonstrate that a magellan (mgn) mutant causes an egg polarity defect and is characterized by an enlarged Bb and a mis-localized nucleus. The mgn mutant specifically, disrupts the microtubule actin crosslinking factor (macf1a) gene, known to function in other polarized cells. The goal of the proposed study will focus on the earliest stages of oogenesis that result in a mature and viable oocyte for successful fertilization. To address Bb disassembly and the interaction with MACF1a this study will 1) employ Next-Generation Sequencing (NGS) technology to measure differential expression of transcripts in macf1a stage-specific oocytes, and 2) visualize MACF1a protein in combination with the Bb at the ultrastructural level. The proposed project will uncover for the first time the transcriptome of a zebrafish maternal effect mutant (macf1) which may lead to greater insight into the genes controlling polarity while providing high-resolution characterization of the MACF1 protein/Balbiani body.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.