Sphingosine kinase 1 is up-regulated during hypoxia in U87MG glioma cells. Role of hypoxia-inducible factors 1 and 2.
Sphingosine 1-phosphate (S1P), a sphingolipid metabolite that plays an important role in the regulation of cell survival, growth, migration, and angiogenesis, acts both inside the cells and as an extracellular mediator through binding to five G protein-coupled receptors (S1P(1-5)). Sphingosine kinase 1 (SK1), the enzyme responsible for S1P production, is overexpressed in many solid tumors, including gliomas. One common feature of these tumors is the presence of "hypoxic regions," characterized by cells expressing high levels of hypoxia-inducible factors HIF-1alpha and HIF-2alpha, two transcription regulators that modulate the levels of proteins with crucial roles in tumor progression. So far, nothing is known about the role and the regulation of SK1 during tumor-induced hypoxia or about SK1 regulation and HIFs. Here we investigated the role of HIF-1alpha and HIF-2alpha in the regulation of SK1 during hypoxic stress in glioma-derived U87MG cells. We report that hypoxia increases SK1 mRNA levels, protein expression, and enzyme activity, followed by intracellular S1P production and S1P release. Interestingly, knockdown of HIF-2alpha by small interfering RNA abolished the induction of SK1 and the production of extracellular S1P after CoCl(2) treatment, whereas HIF-1alpha small interfering RNA resulted in an increase of HIF-2alpha and of SK1 protein levels. Moreover, using chromatin immunoprecipitation analysis, we demonstrate that HIF-2alpha binds the SK1 promoter. Functionally, we demonstrate that conditioned medium from hypoxia-treated tumor cells results in neoangiogenesis in human umbilical vein endothelial cells in a S1P receptor-dependent manner. These studies provide evidence of a link between S1P production as a potent angiogenic agent and the hypoxic phenotype observed in many tumors.
Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, South Carolina 29403; Department of Medical Chemistry, Biochemistry, and Biotechnology, University of Milan, Segrate, Milan 20090, Italy.
Viviana
V
Anelli
Department of Medicine, Medical University of South Carolina, Charleston, South Carolina.
Christopher R
CR
Gault
Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, South Carolina 29403.
Amy B
AB
Cheng
Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, South Carolina 29403; Department of Medicine, Medical University of South Carolina, Charleston, South Carolina; Ralph H. Johnson Veterans Affairs Medical Center, Charleston, South Carolina 29401. Electronic address: obeid@musc.edu.
Lina M
LM
Obeid
D051792:Basic Helix-Loop-Helix Proteins / Q000096:biosynthesis*; D001932:Brain Neoplasms / Q000378:metabolism*; D018450:Disease Progression; D015971:Gene Expression Regulation, Enzymologic; D015972:Gene Expression Regulation, Neoplastic; D005910:Glioma / Q000378:metabolism*; D006801:Humans; D051793:Hypoxia-Inducible Factor 1 / Q000096:biosynthesis*; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000096:biosynthesis*; D011401:Promoter Regions, Genetic; D012333:RNA, Messenger / Q000378:metabolism; D034741:RNA, Small Interfering / Q000378:metabolism; D014158:Transcription, Genetic; D015854:Up-Regulation; D000099157:Sphingosine Kinase; D000099162:Endothelial PAS Domain-Containing Protein 1
D051792:Basic Helix-Loop-Helix Proteins; D051793:Hypoxia-Inducible Factor 1; D017853:Phosphotransferases (Alcohol Group Acceptor); D012333:RNA, Messenger; D034741:RNA, Small Interfering; D000099157:Sphingosine Kinase; D000099162:Endothelial PAS Domain-Containing Protein 1