microRNA-184 induces a commitment switch to epidermal differentiation

miR-184 is a highly evolutionary conserved microRNA (miRNA) from fly to human. The importance of miR-184 was underscored by the discovery that point mutations in miR-184 gene led to corneal/lens blinding disease. However, miR-184-related function in vivo remained unclear. Here, we report that the miR-184 knockout mouse model displayed increased p63 expression in line with epidermal hyperplasia, while forced expression of miR-184 by stem/progenitor cells enhanced the Notch pathway and induced epidermal hypoplasia. In line, miR-184 reduced clonogenicity and accelerated differentiation of human epidermal cells. We showed that by directly repressing cytokeratin 15 (K15) and FIH1, miR-184 induces Notch activation and epidermal differentiation. The disease-causing miR-184C57U mutant failed to repress K15 and FIH1 and to induce Notch activation, suggesting a loss-of-function mechanism. Altogether, we propose that, by targeting K15 and FIH1, miR-184 regulates the transition from proliferation to early differentiation, while mis-expression or mutation in miR-184 results in impaired homeostasis

A conditional Pax6 depletion study with no morphological effect on the adult mouse corneal epithelium

This work was supported by the Wellcome Trust (grant 088876/Z/09/Z to JDW, REH and JMC). The funding body had no role in the design of the study, the collection, analysis and interpretation of data or the preparation of the manuscriptPeer reviewedPublisher PD

Comparison of two related lines of tauGFP transgenic mice designed for lineage tracing

Abstract Background The tauGFP reporter fusion protein is produced nearly ubiquitously by the TgTP6.3 transgene in TP6.3 mice and its localisation to microtubules offers some advantages over soluble GFP as a lineage marker. However, TgTP6.3 Tg/Tg homozygotes are not viable and TgTP6.3 Tg/− hemizygotes are smaller than wild-type. TP6.4 mice carry the TgTP6.4 transgene, which was produced with the same construct used to generate TgTP6.3, so we investigated whether TgTP6.4 had any advantages over TgTP6.3. Results Although TgTP6.4 Tg/Tg homozygotes died before weaning, TgTP6.4 Tg/− hemizygotes were viable and fertile and only males were significantly lighter than wild-type. The TgTP6.4 transgene produced the tauGFP fusion protein by the 2-cell stage and it was widely expressed in adults but tauGFP fluorescence was weak or absent in several tissues, including some neural tissues. The TgTP6.4 transgene expression pattern changed over several years of breeding and mosaic transgene expression became increasingly common in all expressing tissues. This mosaicism was used to visualise clonal lineages in the adrenal cortex of TgTP6.4 Tg/− hemizygotes and these were qualitatively and quantitatively comparable to lineages reported previously for other mosaic transgenic mice, X-inactivation mosaics and chimaeras. Mosaicism occurred less frequently in TP6.3 than TP6.4 mice and was only observed in the corneal epithelium and adrenal cortex. Conclusions Mosaic expression makes the TgTP6.4 transgene unsuitable for use as a conventional cell lineage marker but such mosaicism provides a useful system for visualising clonal lineages that arise during development or maintenance of adult tissues. Differences in the occurrence of mosaicism between related transgenic lines, such as that described for lines TP6.3 and TP6.4, might provide a useful system for investigating the mechanism of transgene silencing

CD200 Expression Marks a Population of Quiescent Limbal Epithelial Stem Cells with Holoclone Forming Ability

©2018 The Authors STEM CELLS published by Wiley Periodicals, Inc. on behalf of AlphaMed Press 2018 One of the main challenges in limbal stem cell (LSC) biology and transplantation is the lack of definitive cell surface markers which can be used to identify and enrich viable LSCs. In this study, expression of 361 cell surface proteins was assessed in ex vivo expanded limbal epithelial cells. One marker, CD200 was selected for further characterization based on expression in a small subset of limbal epithelial cells (2.25% ± 0.69%) and reduced expression through consecutive passaging and calcium induced differentiation. CD200 was localized to a small population of cells at the basal layer of the human and mouse limbal epithelium. CD200+cells were slow cycling and contained the majority of side population (SP) and all the holoclone forming progenitors. CD200+ cells displayed higher expression of LSCs markers including PAX6, WNT7A, CDH3, CK14, CK15, and ABCB5 and lower expression of Ki67 when compared to CD200−. Downregulation of CD200 abrogated the ability of limbal epithelial cells to form holoclones, suggesting an important function for CD200 in the maintenance and/or self-renewal of LSCs. A second marker, CD109, which was expressed in 56.29% ± 13.96% of limbal epithelial cells, was also found to co-localize with ΔNp63 in both human and mouse cornea, albeit more abundantly than CD200. CD109 expression decreased slowly through calcium induced cell differentiation and CD109+ cells were characterized by higher expression of Ki67, when compared to CD109− subpopulation. Together our data suggest that CD200 expression marks a quiescent population of LSCs with holoclone forming potential, while CD109 expression is associated with a proliferative progenitor phenotype. Stem Cells 2018;36:1723–1735