CAREER: Understanding the Molecular Mechanisms of Insect Cuticular Chitin Maintenance

昆虫和其他节肢动物有一个坚硬的外骨骼或角质层，它必须在一个称为蜕皮的过程中脱落，以使动物生长。这个NSF CAREER奖旨在通过测试三种可能的机制来研究昆虫角质层形成的机制，从而保护新合成的角质层在蜕皮过程中免受降解，因为旧的角质层被酶分解。虽然昆虫为人类提供了许多好处，但它们也可能对作物造成广泛的损害，并成为导致致命疾病的病原体的载体。因此，研究结果可能会导致创新的害虫和病媒管理战略。所获得的知识将被纳入现有的课程，并将开发涉及昆虫生理学三维可视化模块的主动学习方法，从而增强该项目的教育影响。该项目将为本科生和研究生提供昆虫生理学、生物化学、分子生物学和生物信息学领域的多学科培训。该项目旨在为来自地方院校的弱势和代表性不足的少数民族女本科生提供全面的实验室研究经验和在地区/国家会议上展示其工作的机会，使这些学生能够从事基于STEM的职业。甲壳素是昆虫表皮（外骨骼）、围食基质（肠道衬里）、气管和蛋壳的重要结构成分。昆虫通过蜕皮或蜕皮定期更换旧的角质层以适应生长。在每次蜕皮期间，新的角质层形成，而旧的角质层在脱落之前同时经历几丁质酶的部分降解。值得注意的是，几丁质酶不降解新的表皮几丁质，尽管在原表皮丰富。以前的研究已经证明了蛋白质Knickkopf（Knk）在防止几丁质酶过早降解新角质层中的关键作用，尽管是通过一种未知的机制。这个CAREER项目研究了如何在蜕皮过程中保护新合成的角质层几丁质不受几丁质酶活性的影响，以确保新形成的角质层的完整性。它将测试的假设，几丁质脱乙酰酶发挥了关键作用，在维护角质层的完整性屏蔽几丁质酶，可能与Knk合作。该项目将（1）检查几丁质脱乙酰化对几丁质酶功能的影响，（2）鉴定几丁质脱乙酰化酶功能所需的辅助蛋白，（3）评估几丁质脱乙酰化在肠道围食基质完整性中的重要性。为了更好地了解几丁质脱乙酰酶在维持角质层完整性方面的功能，该研究将采用各种分子生物学、共聚焦显微镜、透射电子显微镜和质谱技术，并对学生进行培训。该奖项反映了NSF的法定使命，并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，被认为值得支持。

Insects and other arthropods have a hard exoskeleton or cuticle, which must be shed in a process called molting for the animal to grow. This NSF CAREER Award aims to investigate mechanisms involved in insect cuticle formation by testing three possible mechanisms whereby the newly synthesized cuticle is protected from degradation during molting as the old cuticle is broken down by enzymes. While insects provide numerous benefits to humankind, they can also inflict extensive damage to crops and serve as vectors for pathogens that cause deadly diseases. Therefore, the research results could lead to innovative pest and vector management strategies. The acquired knowledge will be integrated into existing curricula and active learning approaches involving 3-D visualization modules on insect physiology will be developed, thereby enhancing the educational impact of the project. The project will offer multidisciplinary training to undergraduate and graduate students in the fields of insect physiology, biochemistry, molecular biology, and bioinformatics. By providing underprivileged and underrepresented minority female undergraduate students from local institutions with comprehensive laboratory research experiences and opportunities to present their work at regional/national conferences, the project seeks to empower those students to pursue STEM-based careers.Chitin serves as a crucial structural component of the insect cuticle (exoskeleton), peritrophic matrix (gut lining), trachea, and eggshell. Insects periodically replace their old cuticle through molting or ecdysis to accommodate growth. During each molt, a new cuticle forms, while the old one simultaneously undergoes partial degradation by chitinases before being shed. Notably, chitinases do not degrade the new cuticular chitin despite being abundant in the procuticle. Previous research has demonstrated the critical role of the protein Knickkopf (Knk) in preventing premature degradation of the new cuticle by chitinases, albeit through an unknown mechanism. This CAREER project investigates how newly synthesized cuticular chitin is shielded from chitinase activity during molting to ensure the integrity of the newly forming cuticle. It will test the hypothesis that chitin deacetylases play a pivotal role in maintaining cuticle integrity by shielding against chitinases, potentially in collaboration with Knk. The project will (1) examine the impact of chitin deacetylation on chitinase function, (2) identify accessory proteins required for chitin deacetylase function, and (3) assess the importance of chitin deacetylation in gut peritrophic matrix integrity. To better understand the function of chitin deacetylases in maintaining cuticle integrity, the research will employ, and train students in, various molecular biology, confocal microscopy, transmission electron microscopy and mass-spectrometry techniques.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.