Analysis of the hypoxia-sensing pathway in Drosophila melanogaster.

摘要: The mechanism by which hypoxia induces gene transcription involves the inhibition of HIF-1α (hypoxia-inducible factor-1 α subunit) PHD (prolyl hydroxylase) activity, prevents VHL (von Hippel-Lindau)-dependent targeting to ubiquitin/proteasome pathway. thus accumulates and promotes transcription. In present study, first we provide direct biochemical evidence for presence a conserved hypoxic signalling pathway in Drosophila melanogaster. An assay 2-oxoglutarate-dependent dioxygenases was developed using embryonic larval homogenates as source enzyme. has low substrate specificity hydroxylates key proline residues ODD (oxygen-dependent degradation) domains human Similar, homologue HIF-1α. enzyme [35S]VHL binding GST (glutathione S-transferase)–ODD-domain fusion protein. Hydroxylation is enhanced proteasomal inhibitors ascertained an anti-hydroxyproline antibody. Secondly, transgenic flies expressing protein that combined domain green fluorescent (ODD–GFP), analysed cascade different tissues. Hypoxic accumulation reporter observed whole tracheal tree, but not ectoderm. stabilization ODD–GFP ectoderm restored inducing expression these cells. These results show tissues exhibit sensitivities hypoxia.