Generation and characterization of a Dkk4-Cre knock-in mouse line.

Ectodermal appendages in mammals, such as teeth, mammary glands, sweat glands and hair follicles, are generated during embryogenesis through a series of mesenchymal-epithelial interactions. Canonical Wnt signaling and its inhibitors are implicated in the early steps of ectodermal appendage development and patterning. To study the activation dynamics of the Wnt target and inhibitor Dickkopf4 (Dkk4) in ectodermal appendages, we used CRSIPR/Cas9 to generate a Dkk4-Cre knock-in mouse (Mus musculus) line, where the Cre recombinase cDNA replaces the expression of endogenous Dkk4. Using Cre reporters, the Dkk4-Cre activity was evident at the prospective sites of ectodermal appendages, overlapping with the Dkk4 mRNA expression. Unexpectedly, a predominantly mesenchymal cell population in the embryo posterior also showed Dkk4-Cre activity. Lineage-tracing suggested that these cells are likely derived from a few Dkk4-Cre-expressing cells in the epiblast at early gastrulation. Finally, our analyses of Dkk4-Cre-expressing cells in developing hair follicle epithelial placodes revealed intra- and inter-placodal cellular heterogeneity, supporting emerging data on the positional and transcriptional cellular variability in placodes. Collectively, we propose the new Dkk4-Cre knock-in mouse line as a suitable model to study Wnt and DKK4 inhibitor dynamics in early mouse development and ectodermal appendage morphogenesis.

High proliferation and delamination during skin epidermal stratification.

The development of complex stratified epithelial barriers in mammals is initiated from single-layered epithelia. How stratification is initiated and fueled are still open questions. Previous studies on skin epidermal stratification suggested a central role for perpendicular/asymmetric cell division orientation of the basal keratinocyte progenitors. Here, we use centrosomes, that organize the mitotic spindle, to test whether cell division orientation and stratification are linked. Genetically ablating centrosomes from the developing epidermis leads to the activation of the p53-, 53BP1- and USP28-dependent mitotic surveillance pathway causing a thinner epidermis and hair follicle arrest. The centrosome/p53-double mutant keratinocyte progenitors significantly alter their division orientation in the later stages without majorly affecting epidermal differentiation. Together with time-lapse imaging and tissue growth dynamics measurements, the data suggest that the first and major phase of epidermal development is boosted by high proliferation rates in both basal and suprabasally-committed keratinocytes as well as cell delamination, whereas the second phase maybe uncoupled from the division orientation of the basal progenitors. The data provide insights for tissue homeostasis and hyperproliferative diseases that may recapitulate developmental programs.