Oxygen sensing in Drosophila: multiple isoforms of the prolyl hydroxylase fatiga have different capacity to regulate HIFalpha/Sima.
BACKGROUND
The Hypoxia Inducible Factor (HIF) mediates cellular adaptations to low oxygen. Prolyl-4-hydroxylases are oxygen sensors that hydroxylate the HIF alpha-subunit, promoting its proteasomal degradation in normoxia. Three HIF-prolyl hydroxylases, encoded by independent genes, PHD1, PHD2, and PHD3, occur in mammals. PHD2, the longest PHD isoform includes a MYND domain, whose biochemical function is unclear. PHD2 and PHD3 genes are induced in hypoxia to shut down HIF dependent transcription upon reoxygenation, while expression of PHD1 is oxygen-independent. The physiologic significance of the diversity of the PHD oxygen sensors is intriguing.


RESULTS
We have analyzed the Drosophila PHD locus, fatiga, which encodes 3 isoforms, FgaA, FgaB and FgaC that are originated through a combination of alternative initiation of transcription and alternative splicing. FgaA includes a MYND domain and is homologous to PHD2, while FgaB and FgaC are shorter isoforms most similar to PHD3. Through a combination of genetic experiments in vivo and molecular analyses in cell culture, we show that fgaB but not fgaA is induced in hypoxia, in a Sima-dependent manner, through a HIF-Responsive Element localized in the first intron of fgaA. The regulatory capacity of FgaB is stronger than that of FgaA, as complete reversion of fga loss-of-function phenotypes is observed upon transgenic expression of the former, and only partial rescue occurs after expression of the latter.


CONCLUSIONS
Diversity of PHD isoforms is a conserved feature in evolution. As in mammals, there are hypoxia-inducible and non-inducible Drosophila PHDs, and a fly isoform including a MYND domain co-exists with isoforms lacking this domain. Our results suggest that the isoform devoid of a MYND domain has stronger regulatory capacity than that including this domain.
Acevedo
Julieta M
JM
Instituto Leloir, Buenos Aires, Argentina.
Centanin
Lazaro
L
Dekanty
Andrés
A
Wappner
Pablo
P
D017398:Alternative Splicing; D000818:Animals; D004268:DNA-Binding Proteins / Q000378:metabolism*; D029721:Drosophila Proteins / Q000378:metabolism*; D004331:Drosophila melanogaster / Q000201:enzymology / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D015971:Gene Expression Regulation, Enzymologic; D056426:Genetic Loci / Q000235:genetics; D006801:Humans; D000860:Hypoxia / Q000378:metabolism; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008018:Life Cycle Stages / Q000235:genetics; D010100:Oxygen / Q000378:metabolism*; D011393:Procollagen-Proline Dioxygenase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000235:genetics; D020218:Response Elements / Q000235:genetics; D015854:Up-Regulation
D004268:DNA-Binding Proteins; D029721:Drosophila Proteins; D007527:Isoenzymes; D012333:RNA, Messenger; C100429:Sima protein, Drosophila; D011393:Procollagen-Proline Dioxygenase; D010100:Oxygen
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