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Fig. 5. The expression of differentiation cardiac markers is downregulated upon loss of Dkk1 and can be rescued by co-injection of Dkk1 MO and human DKK1. A. Front view embryos of stage 28 showing reduced expression of cardiac marker genes on MO injected side (black arrow). Control MO or Dkk1 MO was injected unilaterally at the dorsal-vegetal site in embryos at stage 4. Un-injected side served as internal control. Data quantification is shown in B. C. Co-injection of Dkk1 MO and human DKK1 restore the reduced expression (black arrow) of markers as shown in stage 28 embryos of front view. Data quantification is presented in D. White dotted lines indicate the cement gland. Mean values with standard errors are given. n = number of independent experiments, N = total number of embryos analyzed. *p < 0.05, **p < 0.01 with Mann-Whitney rank sum test. |
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Fig. 7. Wnt/β-catenin activity is increased in the developing heart and inhibition of Wnt/β-catenin signaling rescues loss of Dkk1 phenotype. A. Wnt/β-catenin activity was monitored in the developing heart tissue (marked in pink) isolated from embryos at indicated stages. B. Schematic representation of EnR-LefδN-GR755A construct. Upon injection of this construct into embryos, the expression of LefδN can be induced to inhibit Wnt/β-catenin signaling by adding dexamethasone. C. Expression of cardiac differentiation markers was analyzed in embryos at stage 28 (black arrow). Dotted lines indicate the cement gland. Inhibition of Wnt/β-catenin signaling from stage 20 rescued loss of Dkk1 phenotype in unilaterally injected embryos. n = number of independent experiments, N = total number of embryos analyzed. Mean values with standard errors are given. *p < 0.05, **p < 0.01 with Mann-Whitney rank sum test. |
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Fig. 8. Wnt11a signaling acts downstream of Dkk1 during early cardiogenesis. A. Front view of stage 28 embryos showing the reduced expression of wnt11a and alcam upon unilateral Dkk1 knockdown (black arrow). White dotted lines indicate the cement gland. B. Co-injection of Dkk1 MO with alcam restored the expression of cardiac marker genes (black arrow) at stage 28. C. Co-injection of Dkk1 MO with wnt11a rescued the expression of cardiac marker genes (black arrow) at stage 28. D. Reduced expression of wnt11a by Dkk1 MO injection is restored by inhibiting Wnt/β-catenin signaling. Quantifications are shown next to the image of the embryos. Black dotted lines indicate the cement gland. n = number of independent experiments, N = total number of embryos analyzed. Mean values with standard errors are given. *p < 0.05, **p < 0.01 with Mann-Whitney rank sum test. |
