Figure 4: TF Identification and Validation

(A) Transcription factor (TF) motifs enriched at active promoters (marked by H3K4me3) of differentially expressed genes for each aHCF and fHCF. Expression of each corresponding TF gene taken from RNA-seq data is also shown. (B) Effective >90% GapmeR-mediated knockdown (k.d.) of PLAGL1 and STAT1 in aHCFs shown by reverse transcription quantitative–polymerase chain reaction, normalized to GAPDH and PPIA housekeeping gene control and subsequently to negative A (NegA) control. Values are mean ± SD. (C) Knockdown of either PLAGL1 or STAT1 leads to changes in the aHCF phenotype compared with control specimens. (D) Quantification of cellular parameters after knockdown of either TF shows that aHCFs become smaller and rounder after STAT1 knockdown. PLAGL1 knockdown also results in increased proliferation. All experiments were conducted as n = 3. Biological repeats in independent human cardiac fibroblast cell isolates are shown in Supplemental Figure 6. Values are mean ± SD. *p < 0.05 compared with NegA control. Abbreviations as shown in Figures 1 and 3.