Fig. 5
From: Regulation of MRTF-A by JMY via a nucleation-independent mechanism
Isolated WH2/V domains of JMY activate MRTF-SRF-mediated transcription. Serum-starved NIH 3 T3 cells expressing the isolated V or VVVCA domains of JMY were analyzed for MRTF-SRF activation. a: Schematic overview of generated JMY-V/VVVCA constructs as GFP/GST fusion proteins with domain description. b: Overexpression of isolated JMY-V/VVVCAs detected by immunoblotting with a GFP-specific antibody. Tubulin was used as loading control. c: GFP-JMY-V/VVVCAs and Thymosin β4 induce SRF reporter activity compared to the starved GFP control (GFP). GFP-expressing control cells were stimulated with 15% serum for 7 h (GFP + FCS). d: Endogenous Acta2 mRNA level in serum-starved NIH 3 T3 cells expressing the indicated JMY regions compared to the control (GFP). e: Subcellular localization of endogenous MRTF-A in GFP-JMY-V/VVVCA overexpressing cells. Arrows indicate GFP-positive cells. Scale bars, 20 μm. f: Quantification of E with 50 GFP-positive cells per construct in each of three independent experiments. All data were normalized to the serum-starved GFP control which is set to 1 in c and d. Error bars, s.e.m., n ≥ 3 (* p ≤ 0.05, ** p ≤ 0.01, *** p ≤ 0.001 according to an unpaired two sample student’s t-test (f) or an unpaired one sample student’s t-test (c, d))