Identification of miR-141 as a new regulator of epidermal homeostatis, in cooperation with miR-203

MicroRNAs, small endogenous non-coding RNAs, are involved in the regulation of epidermal homeostasis. Among them, miR-203 was the most described and expressed in human epidermis, promoting keratinocyte differentiation by repressing genes involved in proliferation. To identify other miRNAs involved in this process, the miRNomes of normal human keratinocytes cultured in monolayer (2D) or in 3D reconstructed skin were compared. Besides miR-203, mR-141 was one of the most expressed miRNAs in 3D culture and was overexpressed in 3D vs 2D condition, i.e. during keratinocyte differentiation. Functional experiments revealed that, mostly expressed in the basal layer, miR-141 decreased keratinocyte proliferation and clonogenicity while promoting their differentiation. Target prediction algorithm coupled with transcriptomic data of keratinocytes overexpressing miR-141, as well as 3’UTR luciferase assays enabled to evidence CCND2 mRNA as a direct target of miR-141, leading to its down regulation by miR-141 overexpression. Finally, CCND2 silencing decreased keratinocyte proliferation and induced their differentiation, revealing that miR-141 action was mediated by CCND2. MiR-141 features were also compared with miR-203 in parallel experiments. Although miR-141 displayed similar functions to the ones of miR-203, it exhibited different localization and targets, suggesting a joint participation of miR-141 and miR-203 to engage and maintain keratinocyte towards differentiation, respectively. To discover microRNAs which could be involved in the proliferation / differentiation balance of epidermal keratinocytes, we determined differentially expressed miRNA between normal primary human keratinocytes (NHK) grown in 2D culture (monolayer, n=3 biological replicates) and NHK grown in 3D culture (reconstructed epidermis, 3 biological replicates).