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. 2014 May 23;32(6):1515–1526. doi: 10.1002/stem.1695

Figure 3.

Figure 3

The Sox17-dependent genes Hhex and Cer1 are important for cardiac myogenesis in differentiating ESCs. (A–C): Hhex and (D) Cer1 shRNA suppressed the respective cognate genes in differentiating ESCs and inhibited the induction of cardiac transcription factors and structural genes, acting at a stage subsequent to induction of Mesp1/2. *, p < .05 versus control cells; n ≥ 3. (A, D): Results are shown for the most potent of the shRNAs tested, measured by effectiveness against the endogenous transcripts. For each gene, qualitatively similar results were obtained using at least two independent shRNAs. (B): Partial comparison of the microarray findings with Hhex and Sox17 shRNAs, illustrating the shared impairment of Cer1, cardiac transcription factors, and cardiac structural genes. In addition, a potential positive feedback loop between Hhex and Sox17 is noted. n = 2 for Hhex shRNA; n = 1 for the Luc shRNA controls. (C): Ectopic Cer1 expression rescues cardiac differentiation in Hhex-knock down ESCs. Cer1 was encoded by a tetO-regulated lentiviral vector, and was induced on day 3 by doxycycline. Gene expression was assayed by QRT-PCR. *, p < .05 versus control cells; n ≥ 3.