MKL1 effect on brown/white adipocyte differentiation is not mediated through SRF but rather by regulating PPARγ transcriptional activity.

(A-B) Relative mRNA expression of typical SRF target genes upon siRNA-mediated knockdown of SRF and MKL1 in (A) brown and (B) white immortalized adipocytes. Data are shown as mean + SD, n = 6–8. (C) Luciferase activity from an SRE-Luc construct upon SRF and Mkl1 knockdown. Data are shown as mean + SD, n = 8. (D) Treatment of brown and white pre-adipocytes with indicated concentrations of CCG-1423 from day -2 to day 2 of differentiation. Evaluation of percentage of differentiated cells (left panel) and UCP1-positive cells (right panel). Data are shown as mean ± SD, n = 6. (E) Upregulation of typical PPARγ target genes upon MKL1 knockdown. Data are shown as mean + SD, n = 6. (F) Luciferase activity from a PPRE-containing reporter upon Mkl1 knockdown. Data are shown as mean + SD, n = 6. (G) Co-immunoprecipitation of MKL1 and PPARγ from nuclear extracts of immortalized brown adipocytes on day 4 of differentiation. (H) Co-immunoprecipitation of MKL1 and PPARγ from nuclear extracts of 3T3-L1 adipocytes on day 4 of differentiation. * denotes p-value <0.05, ** denotes p-value <0.01 and *** denotes p-value <0.005 vs. control.