Canonical Wnt signaling regulates Foxc1/2 expression in P19 cells

FOXC1 and FOXC2 are forkhead/winged-helix transcription factors expressed in paraxial mesoderm and somites. Emphasizing the importance of FOXC1/2 during embryonic development, double-knockout mice lacking the alleles for both Foxc1 and Foxc2 failed to form segmented somites and undergo myogenesis. The present study aims to determine upstream factors that regulate Foxc1/2 expression during the differentiation of P19 cells into skeletal muscle. Previous work had shown that dominant-negative forms of beta-catenin, Gli2, and Meox1 could inhibit distinct stages of skeletal myogenesis in P19 cells. In the presence of a dominant-negative beta-catenin fusion protein, Foxc1/2 transcripts were not upregulated and neither were markers of somitogenesis/myogenesis, including Meox1, Pax3 and MyoD. Conversely, inhibition of GSK3 by LiCl or overexpression of activated beta-catenin in aggregated P19 cells resulted in enhancement of Foxc1/2 expression, indicating that FOX transcription may be under the control of Wnt signaling. Supporting this hypothesis, beta-catenin bound to conserved regions upstream of Foxc1 during P19 cell differentiation and drove transcription from this region in a promoter assay. In addition, ectopic expression of a dominant-negative Meox1 or Gli2 resulted in decreased Foxc1/2 transcript levels, correlating with inhibition of skeletal myogenesis. Overexpression of Gli2 was also sufficient to upregulate Foxc1/2 transcript levels and induce skeletal myogenesis. In summary, Foxc1/2 expression is dependent on a complex interplay from various signaling inputs from the Wnt and Shh pathways during early stages of in vitro skeletal myogenesis. Crown Copyright (C) 2009 Published by Elsevier Ltd. on behalf of International Society of Differentiation All rights reserved.