PRFB FY 2021: Characterizing the symbiosome membrane in two divergent cnidarian lineages to investigate the cellular mechanisms and evolution of coral-algal symbiosis

该行动资助了2022财年的NSF生物学博士后研究奖学金，即调查基因组，环境和表型之间相互作用的生命规则的综合研究。该研究金支持研究员的研究和培训，以创新的方式为生活规则领域做出贡献。该研究员将使用尖端的基因编辑和分子技术来研究珊瑚和与珊瑚共生的藻类之间共生的基本机制，这是珊瑚健康的一个关键方面。气候变化威胁到这种共生关系的稳定性，因此可能影响珊瑚的生存。因此，更好地了解这种关系可能有助于制定珊瑚生存的解决方案。该研究员将通过在马里兰州巴尔的摩的外展活动中展示他们的工作来支持科学的多样性和包容性，为当地K-12学生开发使用海葵的实践课程，并为俄勒冈州州立大学的无脊椎动物生物学实验室设计基于研究的课程。蛋白质定位于共生体膜以维持共生，并且其中一些蛋白质在独立进化光共生的宿主之间共享。该研究员将表征共生体膜蛋白质组在两个不同的刺胞动物谱系，独立获得光共生：海葵Exaiptasia diaphana和珊瑚Acropora millepora。为了表征共生体膜蛋白质组，首先，将从每个宿主物种中分离藻细胞，并用抗体标记和流式细胞术富集共生体阳性藻细胞。然后，共生体膜蛋白将纯化与生化和鉴定使用串联质谱。然后，该研究员将通过在两种宿主物种中用CRISPR/Cas9敲除每个基因来测试共生体膜蛋白的功能。将用藻类共生体接种敲除幼虫，并测量共生体密度。预计对共生至关重要的基因的敲除对共生体密度有很大的影响，而那些不太重要的基因的影响则较弱。最后，该研究员将在俄勒冈州州立大学开展基于课程的本科生研究经验，学生将描述E.在共生的早期阶段，diaphana幼虫通过一个时间进程。预期这些基因的表达根据其功能随时间变化。该研究员将接受赞助商在生物信息学、蛋白质组学、CRISPR/cas9敲除技术和课程开发方面的培训。该奖项反映了NSF的法定使命，并被认为值得通过使用基金会的智力价值和更广泛的影响审查标准进行评估来支持。

This action funds an NSF Postdoctoral Research Fellowship in Biology for FY 2022, Integrative Research Investigating the Rules of Life Governing Interactions Between Genomes, Environment and Phenotypes. The fellowship supports research and training of the fellow that will contribute to the area of Rules of Life in innovative ways. The fellow will use cutting-edge gene-editing and molecular techniques to study the fundamental mechanisms governing the symbiosis between corals and algae that live with the coral, which is a key aspect of coral health. Climate change threatens the stability of this symbiosis and therefore can impact coral survival. Better understanding this relationship therefore may help with the development of solutions for coral survival. The fellow will support diversity and inclusion in the sciences by presenting their work at outreach events in Baltimore, MD, developing a hands-on curriculum for local K-12 students using a sea anemone, and by designing a research-based course for Oregon State University’s Invertebrate Biology lab.In cnidarian-algal photosymbiosis, the algal symbiont resides within a phagosomal membrane, called the symbiosome. Proteins localize to the symbiosome membrane to maintain symbiosis, and several of these proteins are shared among hosts that evolved photosymbiosis independently. The fellow will characterize the symbiosome membrane proteome in two divergent cnidarian lineages that have independently gained photosymbiosis: the sea anemone Exaiptasia diaphana and the coral Acropora millepora. To characterize the symbiosome membrane proteome, first, algal cells will be isolated from each host species and symbiosome-positive algal cells will be enriched with antibody labeling and flow cytometry. Then, symbiosome membrane proteins will be purified with biochemistry and identified using tandem mass spectrometry. The fellow will then test the function of symbiosome membrane proteins by knocking out each gene with CRISPR/Cas9 in the two host species. Knockout larvae will be inoculated with algal symbionts and symbiont density will be measured. Knockouts of genes critical for symbiosis are predicted to have a strong effect on symbiont density, while those that are less vital will have weaker effects. Finally, the fellow will develop a course-based undergraduate research experience at Oregon State University where students will characterize the gene expression of symbiosome membrane proteins in E. diaphana larvae through a time course during the early stages of symbiosis. Expression of these genes are expected to vary temporally depending on their function. The fellow will be trained by his sponsors in bioinformatics, proteomics, CRISPR/cas9 knockout techniques, and curriculum development.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.