Molecular imaging: into in vivo interaction of HIF-1alpha and HIF-2alpha with ARNT.
Fluorescence resonance energy transfer (FRET) combined with confocal laser microscopy is a powerful tool to analyze protein-protein interaction in vivo. We have applied this combination to study the assembly of the hypoxia-inducible factor (HIF) complex in living cells under hypoxic conditions. In hypoxia, the basic helix-loop-helix/Period/ARNT/Single-minded (PAS) proteins HIF-1alpha and ... HIF-2alpha accumulate and are translocated into the nucleus. Here, HIF-1alpha and HIF-2alpha dimerize with HIF-1beta, also known as aryl hydrocarbon receptor nuclear translocator (ARNT), to form HIF-1/HIF-2 complexes, which control the expression of specific target genes. Therefore, a new Java-based analyzing program was developed at our institute to calculate the nanometer distance between alpha and beta subunits of the transcriptionally active HIF-1/-2 complex bound to DNA. Fusion proteins of HIF subunits and variants of green fluorescent proteins (cyan and yellow fluorescent proteins) were expressed in living cells and protein-protein interactions were imaged in vivo by means of FRET.
Mesh Terms:
Animals, Aryl Hydrocarbon Receptor Nuclear Translocator, Basic Helix-Loop-Helix Transcription Factors, Cell Line, Tumor, Fluorescence Resonance Energy Transfer, Humans, Hypoxia-Inducible Factor 1, alpha Subunit, Mice, Microscopy, Confocal, Protein Binding, Protein Multimerization
Animals, Aryl Hydrocarbon Receptor Nuclear Translocator, Basic Helix-Loop-Helix Transcription Factors, Cell Line, Tumor, Fluorescence Resonance Energy Transfer, Humans, Hypoxia-Inducible Factor 1, alpha Subunit, Mice, Microscopy, Confocal, Protein Binding, Protein Multimerization
Ann. N. Y. Acad. Sci.
Date: Oct. 01, 2009
PubMed ID: 19845609
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