ThemiR-200family is required for ectodermal organ development through the regulation of the epithelial stem cell niche

Abstract

The murine lower incisor ectodermal organ contains a single epithelial stem cell (SC) niche that provides epithelial progenitor cells to the continuously growing rodent incisor. The dental stem cell niche gives rise to several cell types and we demonstrate that the miR-200 family regulates these cell fates. The miR-200 family is highly enriched in the differentiated dental epithelium and absent in the stem cell niche. In this study, we inhibited the miR-200 family in developing murine embryos using new technology, resulting in an expanded epithelial stem cell niche and lack of cell differentiation. Inhibition of individual miRs within the miR-200 cluster resulted in differential developmental and cell morphology defects. miR-200 inhibition increased the expression of dental epithelial stem cell markers, expanded the stem cell niche and decreased progenitor cell differentiation. RNA-seq. identified miR-200 regulatory pathways involved in cell differentiation and compartmentalization of the stem cell niche. The miR-200 family regulates signaling pathways required for cell differentiation and cell cycle progression. The inhibition of miR-200 decreased the size of the lower incisor due to increased autophagy and cell death. New miR-200 targets demonstrate gene networks and pathways controlling cell differentiation and maintenance of the stem cell niche. This is the first report demonstrating how the miR-200 family is required for in vivo progenitor cell proliferation and differentiation.

Extracellular vesicle expansion of PMIS‐miR‐210 expression inhibits colorectal tumour growth via apoptosis and an XIST/NME1 regulatory mechanism

“…The PMIS inhibitor is composed of native, unmodified nucleic acid that enables the development of stably expressing cells (lentivirus) and animal models for the study of genome‐wide miR family inhibition to identify miR targets and cellular processes. 15 , 16 , 20 , 60 The PMIS inhibitor can be used to knockdown miRs during embryonic development to determine their effect on stem cells, cell proliferation and differentiation as well as developmental processes. 15 , 16 , 17 , 18 , 19 , 20 , 60 The PMIS system allows researchers to finally dissect the role of miRs expressed in clusters and on multiple chromosomes.…”

“…The plasmid‐based microRNA inhibitor system (PMIS) is an miR inhibition system that comprises a 122nt miR binding structure designed to interact with RNA‐induced silencing complex (RISC) factors to load the mature miR into a stable inhibitor complex with high efficiency, specificity and stability 15,16 . This inhibitor is expressed from a plasmid or construct without chemical modifications that produce continual expression of a specific antisense RNA molecule in cells and transgenic mice 15–20 . The PMIS delivers physiological levels of specific miR inhibitors to cells and in transgenic mice with high specificity and no off‐target effects or toxicity 15,16 …”

“… 15 , 16 This inhibitor is expressed from a plasmid or construct without chemical modifications that produce continual expression of a specific antisense RNA molecule in cells and transgenic mice. 15 , 16 , 17 , 18 , 19 , 20 The PMIS delivers physiological levels of specific miR inhibitors to cells and in transgenic mice with high specificity and no off‐target effects or toxicity. 15 , 16 …”

Revisiting the miR-200 Family: A Clan of Five Siblings with Essential Roles in Development and Disease

“…Therefore, the maintenance and differentiation of the lower incisor epithelial population offers an ideal model system to investigate the underlying role of miR-200s in epithelial stem cell renewal and differentiation. Along this line, recent histological and RNA sequencing analyses of murine dental epithelial stem cells show that miR-200 expression is required for the differentiation of terminally located progenitor cells and ameloblasts, as well as during the maintenance of the epithelial dental stem cell niche [ 56 , 57 , 58 ]. Furthermore, embryos depleted of miR-200c show defects in oral epithelial tissue invagination, leading to a reduction in incisor tissue composition and length.…”

“…In particular, miR-200-mediated gene expression regulation is essential during mammalian skin, hair follicle, dental, and sensory organ development. Interestingly, recent studies are beginning to understand the miR-200s’ role in stem cell/progenitor cell populations [ 58 , 230 ], denoting that the role of miR-200s in embryonic and organ development is still underexplored.…”