Fig. 7: Chromatin immunoprecipitation assays highlight the binding sites of EOMES and SOX17 on the PLGF promoter region.

a Schematic showing the putative binding sites of a cardiogenic transcription factor EOMES (5’-[AG]GTGTGA-3’; top) and a vasculogenic transcription factor SOX17 (5’-[C/T]ATTGT[C/G]−3’; bottom) on the human PLGF promoter region. b Chromatin immunoprecipitation (ChIP) assays demonstrated that recruitment of EOMES protein onto one of the putative EOMES-binding motif sites of the human PLGF promoter (−1934 bp upstream from the transcription start site [TSS]) was significantly augmented at days 3 (D3) and 6 (D6) in CM differentiation of WT hESCs. The degree of fold enrichment was larger at day 3 than at day 6. Albeit to a lesser degree, recruitment of EOMES was also detected onto another putative EOMES-binding motif site (−790/−724 bp upstream from the TSS) at D3 and D6. c The ChIP assays demonstrated that recruitment of SOX17 protein was significantly augmented onto one of the putative SOX17-binding motif sites of the human PLGF promoter (−3901 bp upstream from the TSS) at D3 and onto all of the four putative SOX17-binding motif sites (−3901, −1638, −1542, and −1384 bp upstream from the TSS) at D6. Data in b and c are presented as mean ± SD (n = 5 independent experiments). Differences between groups were examined with one-way ANOVA followed by Tukey multiple comparisons test. *P < 0.01 and **P < 0.0001 vs. IgG (negative control). Source data are provided as a Source Data file. d Schematic highlighting the EOMES-PLGF-mediated cardiomyogenesis in the early embryonic stage and the SOX17-PLGF-mediated vasculogenesis in the middle/late embryonic stage. e Schematic highlighting the stage-dependent PLGF’s roles in cardiogenesis. In the early stage, the second heart field (SHF) heart progenitors play a dual role, such as an autocrine role in the SHF and a paracrine role to function to the vascular progenitors (VP). In the late stage, smooth muscle cells (SMCs) play a paracrine role in functioning to endothelial cells (ECs), while ECs play an autocrine role. Black dashed arrows indicate putative differentiation paths. CM cardiomyocyte.