Cloning and expression analysis of two distinct HIF-alpha isoforms--gcHIF-1alpha and gcHIF-4alpha--from the hypoxia-tolerant grass carp, Ctenopharyngodon idellus.

BMC Molecular Biology
Sheran H W LawRichard Y C Kong

Abstract

Hypoxia-inducible factors (HIFs) are involved in adaptive and survival responses to hypoxic stress in mammals. In fish, very little is known about the functions of HIFs. We have cloned and characterized two distinct HIF-alpha cDNAs--gcHIF-1alpha and gcHIF-4alpha--from the hypoxia-tolerant grass carp. The deduced gcHIF-1alpha protein is highly similar to the HIF-1alphas (57-68%) from various vertebrate species, while gcHIF-4alpha is a novel isoform, and shows an equivalent degree of amino acid identity (41-47%) to the HIF-1alpha, HIF-2alpha and HIF-3alpha proteins so far described. Parsimony analysis indicated that gcHIF-4alpha is most closely related to the HIF-3alpha proteins. Northern blot analysis showed that mRNA levels of gcHIF-1alpha and gcHIF-4alpha differ substantially under normoxic and hypoxic conditions, while Western blot studies demonstrated that the endogenous protein levels for both gcHIF-1alpha and gcHIF-4alpha are similarly responsive to hypoxia. Our findings suggest that both gcHIF-1alpha and gcHIF-4alpha are differentially regulated at the transcriptional and translational levels. HRE-luciferase reporter assays show that both proteins function as transcription activators and play distinct roles in modulating ...Continue Reading

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Datasets Mentioned

BETA
AF208487
AB018398
Y09507
AF304864
BC051338
AF402782
AB054067
AAH46875
BX248102
CA471737

Methods Mentioned

BETA
transfection
immunoprecipitation
reverse transcription PCR
PCR

Software Mentioned

PHYLIP
SEQBOOT
BLASTX
BLAST
CONSENSE
TBLASTN
Quantity
PROTPARS
Per

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