Epidermal wound healing is required for normal skin barrier function. Cell motility is a key factor in the ability of keratinocytes to heal epithelial damage. Calmodulin-like protein (CLP) is an epithelial-specific Ca(2+)-binding protein that is regulated during terminal keratinocyte differentiation. CLP is a specific light chain of unconventional myosin-10 (Myo10) and its expression increases filopodial length, filopodial number, and Myo10-dependent cell motility in vitro. However, the effects of CLP expression on keratinocyte motility are unknown. Here we used cultured human keratinocytes to study the role of CLP in regulating Myo10 and the effects of Myo10 and CLP on cell migration. CLP and Myo10 expression were correlated in vitro and required for keratinocyte motility in wound-healing assays. We examined the localization of CLP in wounded skin by immunohistochemistry and found an upregulation and peripheral localization of CLP in the basal and suprabasal cells adjacent to and immediately over the wound bed in vivo. The results suggest that increased CLP expression and CLP-mediated Myo10 function are important for skin differentiation and wound reepithelialization.