Research ArticleCell Biology

Protein kinase A–dependent phosphorylation stimulates the transcriptional activity of hypoxia-inducible factor 1

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Sci. Signal.  31 May 2016:
Vol. 9, Issue 430, pp. ra56
DOI: 10.1126/scisignal.aaf0583

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Enhancing hypoxic responses with PKA

The genes targeted by the transcription factor HIF-1 are involved in adapting to low-oxygen conditions. However, responses mediated by HIF-1 also contribute to the pathogenesis of cancer and cardiovascular disease. The second messenger cAMP is increased in cancer cells and initially increased in failing hearts. Bullen et al. found that the cAMP-activated kinase PKA phosphorylated HIF-1α, which increased its abundance and activity in cultured cardiomyocytes and a cancer cell line. Stimuli that increased cAMP concentrations enhanced the expression of HIF-1 target genes encoding enzymes that convert cAMP to adenosine, a metabolite that suppresses antitumor immunity and reduces heart rate and contractility. Thus, these data establish a mechanistic link between a kinase (PKA) and transcription factor (HIF-1) that contribute to the progression of cancer and cardiovascular disease.

Abstract

Hypoxia-inducible factor 1 (HIF-1) activates the transcription of genes encoding proteins that enable cells to adapt to reduced O2 availability. Proteins encoded by HIF-1 target genes play a central role in mediating physiological processes that are dysregulated in cancer and heart disease. These diseases are also characterized by increased production of cyclic adenosine monophosphate (cAMP), the allosteric activator of cAMP-dependent protein kinase A (PKA). Using glutathione S-transferase pull-down, coimmunoprecipitation, and mass spectrometry analyses, we demonstrated that PKA interacts with HIF-1α in HeLa cervical carcinoma cells and rat cardiomyocytes. PKA phosphorylated Thr63 and Ser692 on HIF-1α in vitro and enhanced HIF transcriptional activity and target gene expression in HeLa cells and rat cardiomyocytes. PKA inhibited the proteasomal degradation of HIF-1α in an O2-independent manner that required the phosphorylation of Thr63 and Ser692 and was not affected by prolyl hydroxylation. PKA also stimulated the binding of the coactivator p300 to HIF-1α to enhance its transcriptional activity and counteracted the inhibitory effect of asparaginyl hydroxylation on the association of p300 with HIF-1α. Furthermore, increased cAMP concentrations enhanced the expression of HIF target genes encoding CD39 and CD73, which are enzymes that convert extracellular adenosine 5′-triphosphate to adenosine, a molecule that enhances tumor immunosuppression and reduces heart rate and contractility. These data link stimuli that promote cAMP signaling, HIF-1α–dependent changes in gene expression, and increased adenosine, all of which contribute to the pathophysiology of cancer and heart disease.

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