The Isl1/Ldb1 Complex Orchestrates Genome-wide Chromatin Organization to Instruct Differentiation of Multipotent Cardiac Progenitors
Ldb1 and Isl1 Interact to Regulate Cardiogenesis
(A) Co-immunoprecipitation of nuclear extracts from d5 EBs using anti-Ldb1 antibody and detected with anti-Isl1 antibody.
(B) Percentage of beating d7 EBs derived from ESCs overexpressing either GFP alone (control) or GFP together with Isl1, Ldb1, or both.
(C–E) Relative mRNA expression of cardiomyocyte (C), endothelial (D), and smooth muscle marker genes (E) in d7 EBs differentiated from ESCs overexpressing either GFP alone or GFP together with Isl1, Ldb1, or Ldb1+Isl1.
(F and G) Relative mRNA expression of cardiac progenitor marker genes in d4 EBs (F) and Flk-1 and PdgfR-α in d3.75 EBs (G) differentiated from ESCs overexpressing either GFP alone or GFP together with Isl1, Ldb1, or Ldb1+Isl1. For qPCR analysis of endogenous Isl1 levels, primers located in the 5′ UTR were utilized.
(H–J) H&E staining of representative paraffin sections of E16.5 hearts of WT and Isl1+/−Ldb1+/− embryos (H, top panels), and higher magnification of right and left ventricles showing thinner compact myocardium of the right ventricle in Isl1+/−Ldb1+/− embryos (H, bottom panels), DORV (I), or OA in E18.5 hearts (J) with VSD (I and J). Ao, Aorta; PA, pulmonary artery; LA, left atrium; LV, left ventricle; RA, right atrium; RV, right ventricle; DORV, double outlet right ventricle; OA, overriding aorta; VSD, ventricular septal defect.
(K) Morphometric analysis of right ventricle compact myocardial thickness. Data are mean ± SEMs, n = 4.
(L) Relative mRNA expression analysis of cardiac progenitor and cardiomyocyte genes in dissected hearts and SHF of E9.25 WT, Isl1+/−, Ldb1+/− (controls), and Isl1+/−Ldb1+/− embryos. Data are mean ± SEMs, n = 4 for each genotype. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.005.
Data in (B)–(G) are mean ± SEMs, n = 3. ∗p < 0.05, ∗∗p < 0.01, ∗∗∗p < 0.005. See also Figure S2 .