Regulation of Drosophila hypoxia-inducible factor (HIF) activity in SL2 cells: Identification of a hypoxia-induced variant isoform of the HIFα homolog gene similar

• Autores: Gorr, T.A.; Tomita, T.; Wappner, P.; Bunn, H.F.
• Año de publicación: 2004
• Idioma: inglés
• Tipo de recurso: artículo
• Fuente: J. Biol. Chem. 2004;279(34):36048-36058
• Materia: Amino acids; Chelation; Genes; Hydroxylation; Iron; Iron chelation; Sima genes; Cell culture; amino acid; cobalt; complementary DNA; deferoxamine; erythropoietin; helix loop helix protein; hypoxia inducible factor 1alpha; iron; luciferase; messenger RNA; metal; oxygen; PAS protein; protein Tango; RNA; Sima protein; unclassified drug; animal cell; article; controlled study; Drosophila; gene overexpression; genetic transfection; hydroxylation; hypoxia; hypoxia response element; molecular cloning; nonhuman; Northern blotting; open reading frame; oxygen tension; priority journal; protein degradation; protein localization; reporter gene; reverse transcription polymerase chain reaction; RNA isolation; signal transduction; transcription regulation; untranslated region; Western blotting; Amino Acid Sequence; Animals; Aryl Hydrocarbon Receptor Nuclear Translocator; Carrier Proteins; Cell Hypoxia; Cell Line; DNA-Binding Proteins; Drosophila; Drosophila Proteins; Gene Expression Regulation; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Molecular Sequence Data; Sequence Homology; Signal Transduction; Transcription Factors; Animalia
• Institución: Universidad Nacional de Buenos Aires. Facultad de Ciencias Exactas y Naturales

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.