Cookies on this website

We use cookies to ensure that we give you the best experience on our website. If you click 'Accept all cookies' we'll assume that you are happy to receive all cookies and you won't see this message again. If you click 'Reject all non-essential cookies' only necessary cookies providing core functionality such as security, network management, and accessibility will be enabled. Click 'Find out more' for information on how to change your cookie settings.

<ns4:p><ns4:bold>Background:</ns4:bold> Little is known about the impact of nutrients on cellular transcriptional responses, especially in face of environmental stressors such as oxygen deprivation. Hypoxia-inducible factors (HIF) coordinate the expression of genes essential for adaptation to oxygen-deprived environments. A second family of oxygen-sensing genes known as the alpha-ketoglutarate-dependent dioxygenases are also implicated in oxygen homeostasis and epigenetic regulation. The relationship between nutritional status and cellular response to hypoxia is understudied. <ns4:italic>In vitro</ns4:italic> cell culture systems frequently propagate cells in media that contains excess nutrients, and this may directly influence transcriptional response in hypoxia.</ns4:p><ns4:p> <ns4:bold>Methods:</ns4:bold> We studied the effect of glucose and glutamine concentration on HepG2 hepatoma transcriptional response to low oxygen and expression of hypoxia inducible factor-1α (HIF-1α). Mass spectrometry confirmed low oxygen perturbation of dioxygenase transcripts resulted in changes in DNA methylation.</ns4:p><ns4:p> <ns4:bold>Results:</ns4:bold> Under normoxic conditions, we observed a significant upregulation of both HIF-target genes and oxygen-dependent dioxygenases in HepG2 cells cultured with physiological levels of glucose or glutamine relative to regular DMEM media, demonstrating that excess glutamine/glucose can mask changes in gene expression. Under hypoxic conditions, <ns4:italic>CA9</ns4:italic> was the most upregulated gene in physiological glutamine media while <ns4:italic>TETs</ns4:italic> and <ns4:italic>FTO</ns4:italic> dioxygenases were downregulated in physiological glucose. Hypoxic regulation of these transcripts did not associate with changes in HIF-1α protein expression. Downregulation of <ns4:italic>TETs</ns4:italic> suggests a potential for epigenetic modulation. Mass-spectrometry quantification of modified DNA bases confirmed our transcript data. Hypoxia resulted in decreased DNA hydroxymethylation, which correlated with <ns4:italic>TETs</ns4:italic> downregulation. Additionally, we observed that <ns4:italic>TET2</ns4:italic> expression was significantly downregulated in patients with hepatocellular carcinoma, suggesting that tumour hypoxia may deregulate <ns4:italic>TET2</ns4:italic> expression resulting in global changes in DNA hydroxymethylation.  </ns4:p><ns4:p> <ns4:bold>Conclusion:</ns4:bold> Given the dramatic effects of nutrient availability on gene expression, future <ns4:italic>in vitro</ns4:italic> experiments should be aware of how excess levels of glutamine and glucose may perturb transcriptional responses.</ns4:p>

Original publication

DOI

10.12688/wellcomeopenres.14839.1

Type

Journal article

Journal

Wellcome Open Research

Publisher

F1000 ( Faculty of 1000 Ltd)

Publication Date

08/10/2018

Volume

3

Pages

126 - 126