CAREER: The Effects of Liquid-Liquid Phase Separation on Ubiquilin Chaperone and Triage Activity

该奖项全部或部分根据2021年美国救援计划法案（公法117-2）资助。为了应对细胞应激，一些蛋白质通过称为液-液相分离（LLPS）的过程形成液体状液滴，但LLPS对蛋白质活性的具体影响知之甚少。其他蛋白质，称为伴侣蛋白，具有减轻细胞应激的作用。这个项目的重点是一个家庭的蛋白质称为Ubiquilins被认为是伴侣。 泛素可以经历LLPS并通过防止有毒蛋白质聚集体的形成来减轻细胞应激。该项目的目标是了解LLPS如何影响Ubiquilins的能力，以帮助蛋白质插入细胞膜或降解不能正确插入的蛋白质，这对于防止有毒蛋白质聚集体的积累和其他形式的细胞应激至关重要。该研究项目与针对寄养儿童和托莱多公立学校（TPS）学生的三项关键外展活动紧密结合。首先，与卢卡斯县儿童服务和TPS合作，将在Elevate夏令营举办科学日，为包括寄养青年在内的高危青年提供安全的夏季环境。营员将学习STEM职业并进行基本的LLPS实验。第二，TPS高中学生将到托莱多大学进行实地考察，参观实验室，揭开大学录取过程的神秘面纱，并了解STEM研究机会。第三，TPS学生和寄养青年将在夏季研究实习指导。这些外展活动将提高科学素养，改善STEM教育，并促进培养青年和TPS学生在STEM领域的充分参与。 美国救援计划基金用于支持这位早期职业研究者在其职业生涯的关键阶段。人类有三种广泛表达的Ubiquilin旁系同源物：Ubqln 1，Ubqln 2和Ubqln 4。已显示泛素结合未插入的膜蛋白，并为生产性膜插入提供额外的机会。泛素还可以与泛素-蛋白酶体系统相互作用以促进客户蛋白质降解。Ubqln 2已被证明经历LLPS，但LLPS如何影响Ubqln 2活性尚不清楚。该项目的重点是确定LLPS如何影响Ubiquilins分子伴侣和分流未插入的膜蛋白的能力。该项目的目标是：（1）确定LLPS如何影响Ubiquilins促进客户蛋白泛素化和降解的能力，以及（2）确定LLPS如何影响Ubiquilins结合膜蛋白并促进其插入脂质双层的能力。该项目的完成将有助于主要研究者的长期目标，即了解LLPS对Ubiquilin活性的功能意义。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知识价值和更广泛的影响审查标准进行评估来支持。

This award is funded in whole or in part under the American Rescue Plan Act of 2021 (Public Law 117-2). In response to cellular stress, some proteins form liquid-like droplets by a process called Liquid-Liquid Phase Separation (LLPS), but the specific effects of LLPS on protein activity are poorly understood. Other proteins, referred to as chaperones, have the role of mitigating cellular stress. This project focuses on a family of proteins called Ubiquilins that are considered chaperones. Ubiquilins can undergo LLPS and mitigate cellular stress by preventing the formation of toxic protein aggregates. The goal of this project is to understand how LLPS affects the ability of Ubiquilins to either assist proteins to insert into the cell membrane or degrade proteins that cannot properly insert, roles that are essential to prevent the accumulation of toxic protein aggregates and other forms of cell stress. The research project is intimately integrated with three key outreach activities targeted to children in foster care and students at Toledo Public Schools (TPS). First, in partnership with Lucas County Children Services and TPS, there will be science day at Elevate Summer Camp, which provides a safe summer environment for at risk youth, including foster youth. Campers will learn about STEM careers and conduct basic LLPS experiments. Second, TPS high school students will do a field trip to the University of Toledo to tour laboratories, demystify the college admission process, and learn about STEM research opportunities. Third, TPS students and foster youth will be mentored in a summer research internship. These outreach activities will increase scientific literacy, improve STEM education, and promote the full participation of foster youth and TPS students in STEM fields. American Rescue Plan funding is used to support this early career investigator at a critical stage in his career.Humans have three broadly expressed Ubiquilin paralogs: Ubqln1, Ubqln2, and Ubqln4. Ubiquilins have been shown to bind to uninserted membrane proteins and provide additional opportunities for productive membrane insertion. Ubiquilins can also interact with the Ubiquitin-Proteasome System to facilitate client protein degradation. Ubqln2 has been shown to undergo LLPS, but how LLPS affects Ubqln2 activity is unknown. This project focuses on determining how LLPS affects the ability of Ubiquilins to chaperone and triage uninserted membrane proteins. The project’s goals are to: (1) determine how LLPS affects the ability of Ubiquilins to facilitate client protein ubiquitination and degradation and (2) determine how LLPS affects the ability of Ubiquilins to bind membrane proteins and facilitate their insertion into the lipid bilayer. Completion of this project will facilitate the principal investigator’s long-term goal of understanding the functional significance of LLPS on Ubiquilin activity.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.