A Foxo/Notch pathway controls myogenic differentiation and fiber type specification
J. Clin. Invest. Tadahiro Kitamura, et al. 117:2477 doi:10.1172/JCI32054 [
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Figure 4Foxo1 binds directly to Csl. (
A) GST pull-down assays of GST-Foxo1 fusion protein with Csl immunoprecipitated from HEK293 cells. (
B and
C) Binding of GST-Foxo1 and GST-FLAG-Csl in a cell-free system and mapping of the Csl interaction domain. Full-length and truncated fragments of GST-Foxo1 and GST-FLAG-Csl were purified from bacteria and coincubated. Thereafter, Csl was isolated using anti-FLAG antibody, and the immunoprecipitate was analyzed by immunoblotting with anti-Foxo1 or anti-FLAG antibodies. (
D)
Hes1 promoter ChIP assay spanning the Csl-binding site in C2C12 cells to detect endogenous Foxo1, Csl, and Notch1 (Endog) or following transduction with Foxo1-ADA during myoblast differentiation. Input represents DNA extracted from chromatin prior to immunoprecipitation.
Hes1 (semiquantitative RT-PCR) and myosin (Western blot) expression corresponding to each time point are shown. Day 0 is defined as the time when cells were serum deprived to induce myoblast fusion.
