1. Author Response:

    We thank the reviewers for their efforts reviewing the manuscript and greatly appreciate the comments and recommendations. We are pleased that the reviewers were in agreement with the main conclusions of the manuscript based on the experimental evidence presented. We are also grateful for the complimentary comments and are encouraged that the reviewers recognized the potential impact of the findings.

    We are thankful for the opportunity to submit a revised manuscript and appreciate the recommendation to include currently missing controls. We agree with the reviewers; our mouse colonies were affected due to long pandemic-related shutdowns, which prevented measurements in all cohorts in a timely fashion. These experiments are now underway, for planned inclusion in the revised manuscript.

    Read the original source
    Was this evaluation helpful?
  2. Reviewer #3 (Public Review):

    The main findings are that loss of the Piezo1 protein in keratinocytes accelerate migration and wound healing, while genetic and pharmacological manipulations known to increase currents carried by Piezo1 slow migration and wound healing. The channels are shown to accumulate and cluster at the trailing edge of single migrating cells and at the wound margin during in vitro studies of wound healing. These findings demonstrate that Piezo1 mechanosensitive channels are not required for keratinocyte migration or wound healing, but rather function as essential regulators of the speed of both migration and would healing. Further, the findings suggest that increased flux through Piezo1 channels slows migration and wound healing. These channels are found to cluster in migrating cells and at wound margins. The conclusions are well-supported by the presented data and the authors' composition does an outstanding job of recognizing the limits of what has been learned and what remains uncertain.

    Read the original source
    Was this evaluation helpful?
  3. Reviewer #2 (Public Review):

    The manuscript "Spatiotemporal dynamics of PIEZO1 localization controls keratinocyte migration during wound healing" by Holt and colleagues demonstrates that loss of function of PIEZO1 speeds up keratinocyte migration and wound closure, whereas enhancing PIEZO1 function, with a PIEZO1 gain-of-function mutant or by chemical means, slows down both processes. The topic of this manuscript is timely and relevant. The experimental design followed by the authors is straightforward and elegant and the vast majority of the conclusions are fully supported by their results. Overall, this manuscript provides solid evidence that normal (wild type) function of PIEZO1 slows down skin wound healing in vitro and in vivo.

    Read the original source
    Was this evaluation helpful?
  4. Reviewer #1 (Public Review):

    In this manuscript, Holt and colleagues investigate how the mechanoreceptor PIEZO1 mediates keratinocyte cell migration and re-epithelialization during wound healing. The authors utilized epidermal-specific Piezo1 knockout mice (Piezo1cKO) and epidermal-specific Piezo1 gain of function mice (Piezo1GoF) to investigate the contribution of keratinocyte Piezo1 to wound healing in vivo. Piezo1cKO mice exhibited faster wound closure, whereas Piezo1GoF mice exhibited slower wound closure compared to controls, suggesting that the presence of epidermal Piezo1 affects the speed of wound healing. To determine if these effects observed in vivo were due to changes in keratinocyte re-epithelization, the authors utilized an in vitro model of wound healing by inducing scratches to mimic "wounds" in keratinocyte monolayers. Similar to the in vivo findings, Piezo1cKO keratinocytes exhibited enhanced wound closure compared to controls. In a separate line of experiments, the authors found that enrichment of Piezo1 at the wound edge induces localized cellular retraction that slows keratinocyte re-epithelization and wound closure. Overall, major strengths are that the topic is of significant interest, Piezo channels and their function is of broad topical interest, and the manuscript is well written. Wound healing is a major health concern and understanding the mechanisms underlying how wounds heal could generate improved therapeutics for faster healing. The key weaknesses are that there are missing controls and missing cohorts (Piezo1GoF or Piezo1cKO) in several of the experimental data sets, and there is a concern about the wide variation in controls for some experiments.

    Read the original source
    Was this evaluation helpful?
  5. Evaluation Summary:

    The manuscript links a critical physiological function of the skin, wound healing to the ability of skin cells to migrate and the modification of migration by the mechanosensitive ion channel Piezo1. The topic of the manuscript is timely, relevant and would be of interest to a broad audience. The experimental design followed by the authors is straightforward and elegant, and the majority of the conclusions are well supported by the results.

    (This preprint has been reviewed by eLife. We include the public reviews from the reviewers here; the authors also receive private feedback with suggested changes to the manuscript. The reviewers remained anonymous to the authors.)

    Read the original source
    Was this evaluation helpful?