A role for the Tgf-β/Bmp co-receptor Endoglin in the molecular oscillator that regulates the hair follicle cycle.

The hair follicle is a biological oscillator that alternates growth, regression, and rest phases driven by the sequential activation of the proliferation/differentiation programs of resident stem cell populations. The activation of hair follicle stem cell niches and subsequent entry into the growing phase is mainly regulated by Wnt/β-catenin signalling, while regression and resting phases are mainly regulated by Tgf-β/Bmp/Smad activity. A major question still unresolved is the nature of the molecular switch that dictates the coordinated transition between both signalling pathways. Here we have focused on the role of Endoglin (Eng), a key co-receptor for members of the Tgf-β/Bmp family of growth factors.Using an Eng haploinsufficient mouse model, we report that Eng is required to maintain a correct follicle cycling pattern and for an adequate stimulation of hair follicle stem cell niches. We further report that β-catenin binds to the Eng promoter depending on Bmp signalling. Moreover, we show that β-catenin interacts with Smad4 in a Bmp/Eng-dependent context and both proteins act synergistically to activate Eng promoter transcription. These observations point to the existence of a growth/rest switching mechanism in the hair follicle that is based on an Eng-dependent feedback crosstalk between Wnt/β-catenin and Bmp/Smad signals. Implication of Endoglin, Wnt/β-catenin and Bmp/Smad signals in the growth/rest hair follicle Microarray experiments were performed using Mouse Gene Expression 4x44K Microarray Kit G4122F (Agilent technologies, Wilmington, DE). RNA was isolated using RNAesy Extraction Kit (QIAGen, Germany). RNA was labeled and array hybridized us ing the Low RNA Linear Amplification Kit and the In Situ Hybridization Kit Plus (Agilent technologies, Wilmington, DE) respectively. After hybridization and washing, the slides were scanned in an Axon GenePix Scanner (Axon Instruments Inc., Union City, CA) and analysed using Feature Extraction Software 10.1 (Agilent technologies, Wilmington, DE). Two different RNA samples obtained from each Eng modified cell line were labelled with Cy5-dUTP. The RNA samples extracted Downloaded from https://academic.oup.com/jmcb/advance-article-abstract/doi/10.1093/jmcb/mjy051/5101434 by Bukkyo University user on 23 September 2018 23from wild type cells were marked with Cy3-dUTP (Amersham, Sweden). Two additional hybridizations were performed using the reciprocal fluorochrome labelling . The genes whose expression was up or downregulated at least 2-fold in Eng+/-with respect to control cells were selected for analysis.