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Theses Doctoral

Characterization of Primary Cilia and Intraflagellar Transport 20 in the Epidermis

Su, Steven

Mammalian skin is a dynamic organ that constantly undergoes self-renewal during homeostasis and regenerates in response to injury. Crucial for the skin’s self-renewal and regenerative capabilities is the epidermis and its stem cell populations. Here we have interrogated the role of primary cilia and Intraflagellar Transport 20 (Ift20) in epidermal development as well as during homeostasis and wound healing in postnatal, adult skin. Using a transgenic mouse model with fluorescent markers for primary cilia and basal bodies, we characterized epidermal primary cilia during embryonic development as well as in postnatal and adult skin and find that both the Interfollicular Epidermis (IFE) and hair follicles (HFs) are highly ciliated throughout development as well as in postnatal and adult skin. Leveraging this transgenic mouse, we also developed a technique for live imaging of epidermal primary cilia in ex vivo mouse embryos and discovered that epidermal primary cilia undergo ectocytosis, a ciliary mechanism previously only observed in vitro. We also generated a mouse model for targeted ablation of Ift20 in the hair follicle stem cells (HF-SCs) of adult mice. We find that loss of Ift20 in HF-SCs inhibits ciliogenesis, as expected, but strikingly it also inhibits hair regrowth. Closer examination of these mice reveals that Ift20 is crucial in maintaining HF-SC identity. Specifically, ablation of Ift20 in HF-SCs results in loss of SOX9 expression in HF-SCs and results in ectopic expression of the IFE marker KLF5 in HF-SCs. Additionally, ectopic differentiation is observed in HF-SCs following loss of Ift20. Finally, using both in vitro and in vivo models, we also characterize the role of primary cilia and Ift20 in epidermal wound healing. We find that loss of Ift20 slows collective keratinocyte migration in vitro and also slows HF-SC migration in vivo during wound repair. Interestingly our data suggests that Ift20 regulates keratinocyte migration in a primary cilia-independent manner. Instead, we find that Ift20 mediates focal adhesion (FA) turnover during keratinocyte migration. Specifically, Ift20 together with Rab5, regulates recycling of FA integrins and loss of Ift20 inhibits proper return of integrins to the keratinocyte surface. Overall, we demonstrate that the epidermis is highly ciliated throughout development and in postnatal skin. We show that Ift20 is crucial in maintaining HF-SC identity and the telogen to anagen transition in HFs. We finally demonstrate that Ift20 regulates keratinocyte migration independent of its function in ciliogenesis and instead regulates recycling of FA integrins through a Rab5 dependent mechanism.

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More About This Work

Academic Units
Cellular, Molecular and Biomedical Studies
Thesis Advisors
Ezratty, Ellen J.
Degree
Ph.D., Columbia University
Published Here
June 29, 2020