Hey bHLH Factors in Cardiovascular Development

Wiese, Cornelia; Heisig, Julia; Gessler, Manfred

doi:10.1007/s00246-009-9609-9

Cited by 23 publications

(23 citation statements)

References 42 publications

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“…Interestingly, it was recently shown that reduced Hey gene expression in endocardial cells leads to impaired EMT of endocardial cells and thus defective cardiac valve formation. [25][26][27] The recent discovery that somatic cells can be first reprogrammed back to pluripotent stem cells and then driven by defined transcriptional cocktails to various cell fates 3 provides a potential avenue for autologous cardiac tissue engineering. We demonstrate here for the first time that PKD2-HDAC5-KLF signaling plays a crucial role in vertebrate AV valve formation.…”

Section: Discussionmentioning

confidence: 99%

“…[25][26][27] To assess whether reduced notch1b expression in bng AV endocardial cells leads to the misexpression of its downstream target genes Hey1, Hey2, and HeyL, we assayed their expression by whole-mount antisense RNA in situ hybridization and cardiac-specific quantitative real-time polymerase chain reaction. We find that hey1 and hey2 are highly expressed at the AV boundary in wild-type embryos.…”

Section: Protein Kinase D2 Controls Atrioventricular Valve Formation mentioning

confidence: 99%

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Protein Kinase D2 Controls Cardiac Valve Formation in Zebrafish by Regulating Histone Deacetylase 5 Activity

et al. 2011

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Background— The molecular mechanisms that guide heart valve formation are not well understood. However, elucidation of the genetic basis of congenital heart disease is one of the prerequisites for the development of tissue-engineered heart valves. Methods and Results— We isolated here a mutation in zebrafish, bungee ( bng jh177 ), which selectively perturbs valve formation in the embryonic heart by abrogating endocardial Notch signaling in cardiac cushions. We found by positional cloning that the bng phenotype is caused by a missense mutation (Y849N) in zebrafish protein kinase D2 ( pkd2 ). The bng mutation selectively impairs PKD2 kinase activity and hence Histone deacetylase 5 phosphorylation, nuclear export, and inactivation. As a result, the expression of Histone deacetylase 5 target genes Krüppel-like factor 2a and 4a, transcription factors known to be pivotal for heart valve formation and to act upstream of Notch signaling, is severely downregulated in bungee ( bng ) mutant embryos. Accordingly, the expression of Notch target genes, such as Hey1, Hey2, and HeyL, is severely decreased in bng mutant embryos. Remarkably, downregulation of Histone deacetylase 5 activity in homozygous bng mutant embryos can rescue the mutant phenotype and reconstitutes notch1b expression in atrioventricular endocardial cells. Conclusions— We demonstrate for the first time that proper heart valve formation critically depends on Protein kinase D2-Histone deacetylase 5-Krüppel-like factor signaling.

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Section: Discussionmentioning

confidence: 99%

Section: Protein Kinase D2 Controls Atrioventricular Valve Formation mentioning

confidence: 99%

Protein Kinase D2 Controls Cardiac Valve Formation in Zebrafish by Regulating Histone Deacetylase 5 Activity

et al. 2011

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“…The intracellular domain of Notch (NICD) is released and translocates into the cell nucleus, where it binds with the CSL complex, leading to activation of target genes such as the Hes/Hey family of transcription factors. [27-29] As such, expression of Hes and Hey genes is used to indicate that Notch signaling is functional in a cell. Our previous study showed that Notch receptors and ligands were expressed in HemSCs and HemECs and we were interested in determining if Notch target genes were expressed in these cells.…”

Section: Discussionmentioning

confidence: 99%

Expression of HES and HEY genes in infantile hemangiomas

Adepoju¹,

Wong²,

Kitajewski³

et al. 2011

Vasc Cell

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BackgroundInfantile hemangiomas (IHs) are the most common benign tumor of infancy, yet their pathogenesis is poorly understood. IHs are believed to originate from a progenitor cell, the hemangioma stem cell (HemSC). Recent studies by our group showed that NOTCH proteins and NOTCH ligands are expressed in hemangiomas, indicating Notch signaling may be active in IHs. We sought to investigate downstream activation of Notch signaling in hemangioma cells by evaluating the expression of the basic HLH family proteins, HES/HEY, in IHs.Materials and MethodsHemSCs and hemangioma endothelial cells (HemECs) are isolated from freshly resected hemangioma specimens. Quantitative RT-PCR was performed to probe for relative gene transcript levels (normalized to beta-actin). Immunofluorescence was performed to evaluate protein expression. Co-localization studies were performed with CD31 (endothelial cells) and NOTCH3 (peri-vascular, non-endothelial cells). HemSCs were treated with the gamma secretase inhibitor (GSI) Compound E, and gene transcript levels were quantified with real-time PCR.ResultsHEY1, HEYL, and HES1 are highly expressed in HemSCs, while HEY2 is highly expressed in HemECs. Protein expression evaluation by immunofluorescence confirms that HEY2 is expressed by HemECs (CD31+ cells), while HEY1, HEYL, and HES1 are more widely expressed and mostly expressed by perivascular cells of hemangiomas. Inhibition of Notch signaling by addition of GSI resulted in down-regulation of HES/HEY genes.ConclusionsHES/HEY genes are expressed in IHs in cell type specific patterns; HEY2 is expressed in HemECs and HEY1, HEYL, HES1 are expressed in HemSCs. This pattern suggests that HEY/HES genes act downstream of Notch receptors that function in distinct cell types of IHs. HES/HEY gene transcripts are decreased with the addition of a gamma-secretase inhibitor, Compound E, demonstrating that Notch signaling is active in infantile hemangioma cells.

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“…NOTCH signaling is a key regulatory pathway for cardiovascular development and homeostasis [1]. Its receptors mainly act through transcriptional activation of target genes by a complex of the NOTCH intracellular domain, released by gamma-secretase, the transcription factor CBF1 (RBP-Jk) and the Mastermind co-activator proteins (Maml1-3).…”

Section: Introductionmentioning

confidence: 99%

Target Gene Analysis by Microarrays and Chromatin Immunoprecipitation Identifies HEY Proteins as Highly Redundant bHLH Repressors

et al. 2012

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HEY bHLH transcription factors have been shown to regulate multiple key steps in cardiovascular development. They can be induced by activated NOTCH receptors, but other upstream stimuli mediated by TGFß and BMP receptors may elicit a similar response. While the basic and helix-loop-helix domains exhibit strong similarity, large parts of the proteins are still unique and may serve divergent functions. The striking overlap of cardiac defects in HEY2 and combined HEY1/HEYL knockout mice suggested that all three HEY genes fulfill overlapping function in target cells. We therefore sought to identify target genes for HEY proteins by microarray expression and ChIPseq analyses in HEK293 cells, cardiomyocytes, and murine hearts. HEY proteins were found to modulate expression of their target gene to a rather limited extent, but with striking functional interchangeability between HEY factors. Chromatin immunoprecipitation revealed a much greater number of potential binding sites that again largely overlap between HEY factors. Binding sites are clustered in the proximal promoter region especially of transcriptional regulators or developmental control genes. Multiple lines of evidence suggest that HEY proteins primarily act as direct transcriptional repressors, while gene activation seems to be due to secondary or indirect effects. Mutagenesis of putative DNA binding residues supports the notion of direct DNA binding. While class B E-box sequences (CACGYG) clearly represent preferred target sequences, there must be additional and more loosely defined modes of DNA binding since many of the target promoters that are efficiently bound by HEY proteins do not contain an E-box motif. These data clearly establish the three HEY bHLH factors as highly redundant transcriptional repressors in vitro and in vivo, which explains the combinatorial action observed in different tissues with overlapping expression.

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Hey bHLH Factors in Cardiovascular Development

Cited by 23 publications

References 42 publications

Protein Kinase D2 Controls Cardiac Valve Formation in Zebrafish by Regulating Histone Deacetylase 5 Activity

Protein Kinase D2 Controls Cardiac Valve Formation in Zebrafish by Regulating Histone Deacetylase 5 Activity

Expression of HES and HEY genes in infantile hemangiomas

Target Gene Analysis by Microarrays and Chromatin Immunoprecipitation Identifies HEY Proteins as Highly Redundant bHLH Repressors

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