miRNA-29-CLIP uncovers new targets and functions to improve skin repair

Abstract

MicroRNAs (miRNAs) control organogenesis in mammals but their role in specific cell types is not fully explored. miRNAs exert their function by binding mRNAs and inhibiting translation. Skin is an excellent model to study the role of miRNAs in primary cells of epidermal (keratinocytes) and mesodermal (fibroblasts) origin, because the growth of these cells is tightly controlled at translation. Previous research demonstrated that miRNA-29 family functions during skin repair, however, the exact mRNA targets and the downstream mechanisms of miRNA-29-mediated regulation of cell growth is missing. Here, we use miRNA crosslinking and immunoprecipitation (miRNA-CLIP) method to find the direct targets of miRNA-29 in keratinocytes and fibroblasts from human skin. We uncover previously unrecognized roles of miRNA-29 in protein folding and RNA processing, common to all cell types tested, and determine the cell-specific role of miRNA-29. Using modified anti-sense oligonucleotides (ASO) in 2D and 3D cultures of keratinocytes and fibroblasts, we enhanced cell-to-matrix adhesion and found an autocrine and paracrine mechanism of miRNA-29-dependent cell growth. Our results include a comprehensive list and functional analyses of mRNAs directly bound by miRNA-29 keratinocytes and fibroblasts, determined by miRNA-CLIP and ASO-mediated inhibition of miRNA-29 followed by RNA-seq. We reveal a full transcriptome of human keratinocytes with enhanced adhesion to the wound matrix, which supports regeneration of the epidermis and is regulated by miRNA-29. The functions of miRNA-29 identified in this study can provide a new approach to improve cutaneous repair by restoring and enhancing the endogenous mechanisms through the stage-specific delivery of miRNA-29 ASO.