Regulation of Drosophila hypoxia-inducible factor (HIF) activity in SL2 cells: identification of a hypoxia-induced variant isoform of the HIFalpha homolog gene similar.
Copyright policy )Regulation of Drosophila hypoxia-inducible factor (HIF) activity in SL2 cells: identification of a hypoxia-induced variant isoform of the HIFalpha homolog gene similar.
Although hypoxia-inducible factor-α (HIFα) subunit-specific hydroxylation and proteolytic breakdown explain the binary switch between the presence (hypoxia) and absence (normoxia) of HIFs, little is known of the mechanisms that fine-tune HIF activity under constant, rather than changing, oxygen tensions. Here, we report that the Drosophila HIFα homolog, the basic helix-loop-helix/PAS protein Sima (Similar), in hypoxic cultures of SL2 cells is expressed in full-length (fl) and splice variant (sv) isoforms. The following evidence supports the role of flSima as functional HIFα and the role of SL2 HIF as a transcriptional activator or suppressor. The pO2 dependence of Sima abundance matched that of HIF activity. HIF-dependent changes in candidate target gene expression were detected through variously effective stimuli: hypoxia (strong) > iron chelation, e.g. desferrioxamine (moderate) ≪ transition metals, e.g. cobalt ≃ normoxia (ineffective). Sima overexpression augmented hypoxic induction or suppression of different targets. In addition to the full-length exon 1-12 transcript yielding the 1510-amino acid HIFα homolog, the sima gene also expressed, specifically under hypoxia, an exon 1-7/12 splice variant, which translated into a 426-amino acid Sima truncation termed svSima. svSima contains basic helix-loop-helix and PAS sequences identical to those of flSima, but, because of deletion of exons 8-11, lacks the oxygen-dependent degradation domain and nuclear localization signals. Overexpressed svSima failed to transactivate reporter genes. However, it attenuated HIF (Sima-Tango)-stimulated reporter expression in a dose-dependent manner. Thus, svSima has the potential to regulate Drosophila HIF function under steady and hypoxic pO2 by creating a cytosolic sink for the Sima partner protein Tango.
Fil:Wappner, P. Universidad de Buenos Aires. Facultad de Ciencias Exactas y Naturales; Argentina.
- Harvard University United States
- Brigham and Women's Faulkner Hospital United States
- University of Buenos Aires Argentina
molecular cloning, PAS protein, oxygen tension, complementary DNA, Sequence Homology, animal cell, hydroxylation, open reading frame, Western blotting, Sima protein, iron, Drosophila Proteins, Sima genes, deferoxamine, Chelation, messenger RNA, article, luciferase, cobalt, reporter gene, Cell Hypoxia, unclassified drug, DNA-Binding Proteins, RNA isolation, priority journal, protein degradation, Amino acids, Iron chelation, Drosophila, erythropoietin, helix loop helix protein, transcription regulation, amino acid, signal transduction, Signal Transduction, metal, Iron, gene overexpression, Molecular Sequence Data, protein localization, Hydroxylation, Cell Line, reverse transcription polymerase chain reaction, untranslated region, Animalia, Animals, Humans, controlled study, hypoxia inducible factor 1alpha, Amino Acid Sequence, nonhuman, hypoxia, Aryl Hydrocarbon Receptor Nuclear Translocator, Hypoxia-Inducible Factor 1, alpha Subunit, protein Tango, Genes, Gene Expression Regulation, RNA, Northern blotting, Cell culture, genetic transfection, Carrier Proteins, oxygen, hypoxia response element, Transcription Factors
molecular cloning, PAS protein, oxygen tension, complementary DNA, Sequence Homology, animal cell, hydroxylation, open reading frame, Western blotting, Sima protein, iron, Drosophila Proteins, Sima genes, deferoxamine, Chelation, messenger RNA, article, luciferase, cobalt, reporter gene, Cell Hypoxia, unclassified drug, DNA-Binding Proteins, RNA isolation, priority journal, protein degradation, Amino acids, Iron chelation, Drosophila, erythropoietin, helix loop helix protein, transcription regulation, amino acid, signal transduction, Signal Transduction, metal, Iron, gene overexpression, Molecular Sequence Data, protein localization, Hydroxylation, Cell Line, reverse transcription polymerase chain reaction, untranslated region, Animalia, Animals, Humans, controlled study, hypoxia inducible factor 1alpha, Amino Acid Sequence, nonhuman, hypoxia, Aryl Hydrocarbon Receptor Nuclear Translocator, Hypoxia-Inducible Factor 1, alpha Subunit, protein Tango, Genes, Gene Expression Regulation, RNA, Northern blotting, Cell culture, genetic transfection, Carrier Proteins, oxygen, hypoxia response element, Transcription Factors
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