(A) NFATc1 mRNA expression was decreased in KLF15 overexpressed podocytes. (B) Consistent with mRNA expression, the protein expression of NFATc1 was reduced in KLF15 overexpressed podocytes. (C) In contrast, silencing of KLF15 elevated NFATc1 mRNA expression in podocytes. (D) Similarly, NFATc1 protein expression was increased in KLF15 silenced podocytes. (E and F) Chormatin immunoprecipitation (ChIP) was performed using an antibody to KLF15, followed by qPCR using the NFATc1 gene promoter specific primer designed at the region -1984 to -1861 base pairs upstream of the transcription start site. DNA electrophoretogram demonstrated that KLF15 bound to NFATc1 promoter region (E) and ChIP-qPCR further demonstrated that the binding amount was decreased in LPS-treated podocytes (F). IgG was used as negative control and input fraction was used as positive control. Fold enrichment = [%(ChIP/Input)]/[%(Negative control/Input). (G) KLF15 directly mediated the regulation of NFATc1 gene promoter activity. Dual-luciferase reporter assay showed that the transcriptional activity of NFATc1 was decreased in KLF15 overexpressed podocytes. The secreted alkaline phosphatase (SEAP) was used as an internal control and Gaussia luciferase (Gluc) was normalized to SEAP, respectively. (H) Fzd9, Rcan1 and plaur, the downstream target genes of NFAT signaling, were decreased in KLF15 overexpressed podocyte. (I) In contrast, silencing of KLF15 increased the mRNA expression of Fzd9, Rcan1 and plaur in podocytes. All data were shown as mean ± SEM. *P<0.05, **P<0.01 and ***P<0.001 vs. controls.