HIF-1alpha signaling upstream of NKX2.5 is required for cardiac development in Xenopus.
HIF-1alpha is originally identified as a transcription factor that activates gene expression in response to hypoxia. In metazoans, HIF-1alpha functions as a master regulator of oxygen homeostasis and regulates adaptive responses to change in oxygen tension during embryogenesis, tissue ischemia, and tumorigenesis. Because Hif-1alpha-deficient mice exhibit a number of developmental defects, the precise role of HIF-1alpha in early cardiac morphogenesis has been uncertain. Therefore, to clarify the role of HIF-1alpha in heart development, we investigated the effect of knockdown of HIF-1alpha in Xenopus embryos using antisense morpholino oligonucleotide microinjection techniques. Knockdown of HIF-1alpha resulted in defects of cardiogenesis. Whole mount in situ hybridization for cardiac troponin I (cTnI) showed the two separated populations of cardiomyocytes, which is indicative of cardia bifida, in HIF-1alpha-depleted embryos. Furthermore, the depletion of HIF-1alpha led to the reduction in cTnI expression, suggesting the correlation between HIF-1alpha and cardiac differentiation. We further examined the expression of several heart markers, nkx2.5, gata4, tbx5, bmp4, hand1, and hand2 in HIF-1alpha-depleted embryos. Among them, the expression of nkx2.5 was significantly reduced. Luciferase reporter assay using the Nkx2.5 promoter showed that knockdown of HIF-1alpha decreased its promoter activity. The cardiac abnormality in the HIF-1alpha-depleted embryo was restored with co-injection of nkx2.5 mRNA. Collectively, these findings reveal that HIF-1alpha-regulated nkx2.5 expression is required for heart development in Xenopus.
Nagao
Kaori
K
Department of Clinical Gene Therapy, Graduate School of Medicine, Osaka University, Suita 565-0871, Japan.
Taniyama
Yoshiaki
Y
Kietzmann
Thomas
T
Doi
Takefumi
T
Komuro
Issei
I
Morishita
Ryuichi
R
D000818:Animals; D005786:Gene Expression Regulation; D018507:Gene Expression Regulation, Developmental; D006321:Heart / Q000196:embryology*; D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins / Q000378:metabolism / Q000502:physiology*; D007150:Immunohistochemistry / Q000379:methods; D017403:In Situ Hybridization; D008954:Models, Biological; D032383:Myocytes, Cardiac / Q000378:metabolism; D009841:Oligonucleotides / Q000737:chemistry; D010100:Oxygen / Q000378:metabolism; D010641:Phenotype; D011401:Promoter Regions, Genetic; D014157:Transcription Factors / Q000378:metabolism / Q000502:physiology*; D019210:Troponin I / Q000378:metabolism; D029867:Xenopus Proteins / Q000378:metabolism / Q000502:physiology*; D014982:Xenopus laevis / Q000196:embryology*
D000072576:Homeobox Protein Nkx-2.5; D018398:Homeodomain Proteins; C092720:NKX2-5 protein, Xenopus; D009841:Oligonucleotides; D014157:Transcription Factors; D019210:Troponin I; D029867:Xenopus Proteins; D010100:Oxygen