An investigation of the roles of mechanical signaling in YAP-mediated tooth renew

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 DESCRIPTION (provided by applicant): Dr. Hu is determined to become an independent researcher to investigate the mechanisms that regulate the morphogenesis and homeostasis of craniofacial structures and to apply that knowledge for translational purposes. CANDIDATE BACKGROUND: Dr. Hu did his graduate research in Dr. Clifford Tabin's laboratory at Harvard University, where he studied limb patterning and muscle development, and gained experiences in developmental biology techniques, mouse genetics, live imaging, and cell culture. Dr. Hu then joined Dr. Ophir Klein's laboratory at UCSF as a postdoctoral fellow to study the mechanisms that regulate mouse incisor stem cells, a great system for understanding stem cell-based renewal. In particular, he found YAP/TAZ as important factors for controlling stem cell proliferation and differentiation and his preliminary data suggest that mechanical signaling plays a key role in regulating YAP activity and stem cell biology. During this time, Dr. Hu gained knowledge in craniofacial and stem cell biology, and has embarked on using the incisor as a system to study mechanotransduction, while developing techniques for culturing, imaging, and measuring force. RESEARCH PLAN: Dr. Hu hypothesizes that nuclear YAP localization and YAP-mediated cellular processes are initially inhibited by actomyosin tension in the incisor stem cells and later upregulatd by integrin/FAK signaling in the transit-amplifying cells. Aim 1 (K99 portion) studies the role of cellular tension by first characterizing cellular features affected by tension, such as cell shapes, levels of phospho-Myosin II, and actin distribution in the stem cell compartment. The effects of tissue tension on cell biology and YAP localization will be examined by laser ablation, and conditional deletion of Myosin IIA/B. Aim 2 (R00 portion) focuses on integrin/FAK signaling. First, the ECM compositions in the stem cell niche will be characterized. Functional studies will then be carried out by testing the ability of ECM proteins t regulate YAP activity and cell differentiation in a 3D culture system, as well as by examining two mouse mutants, one with FAK deletion and the other with a dominant active integrin β1. Finally, a chemical screen will be performed to identify novel factors and pathways that regulate YAP and stem cell biology. Together, these Aims will address key questions in both the Hippo and stem cell field. TRAINING: Dr. Hu will learn several new techniques during the mentored phase, including laser ablation, 3D culturing, mouse knock-ins, and high throughput small molecule screening. He will also attend courses relevant to the study. In addition, Dr. Hu has assembled an advisory committee to help complete and evaluate the project. Finally, he will continue to develop his writing, presentation, managing, and mentoring skills in order to become an independent researcher. ENVIRONMENT: There are many resources and researchers in different fields at UCSF and the greater Bay area science community that are available to Dr. Hu. There are also many regular seminars, workshops, and meetings at UCSF that foster scientific interactions, sharing of ideas, collaborations, and learning opportunities. HEALTH RELATEDNESS: As adult humans don't have dental epithelial stem cells, completion of this study will provide genetic, biomechanical, and chemical targets for developing strategies to derive and maintain clinically safe dental stem cells that can be used to make replacement teeth for treating patients with tooth loss.