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Papers In Press, published online ahead of print August 1, 2001
Genetics, University of Melbourne, Parkville, Victoria 3010
Corresponding Author: m.hynes{at}genetics.unimelb.edu.au
NADP-dependent isocitrate dehydrogenase enzymes catalyze the decarboxylation of isocitrate to 2-oxoglutarate accompanied by the production of NADPH. In mammals two different genes encode mitochondrial and cytoplasmic/peroxisomal located enzymes, while in Saccharomyces cerevisiae three separate genes specify compartment specific enzymes. We have identified a single gene, idpA, in the filamentous fungus Aspergillus nidulans that specifies a protein with a high degree of identity to mammalian and S. cerevisiae enzymes. Northern blot analysis and RT-PCR revealed the presence of two idpA transcripts and two transcription start points were identified by sequencing cDNA clones and by 5'RACE. The shorter transcript was found to be inducible by acetate and by fatty acids while the longer transcript was present in higher amounts during growth in glucose containing media. The longer transcript is predicted to encode a polypeptide containing an N-terminal mitochondrial targetting sequence as well as a C-terminal tripeptide (ARL) as a potential peroxisomal targetting signal. The shorter transcript is predicted to encode a polypeptide lacking the mitochondrial targetting signal but retaining the C-terminal sequence. Immunoblotting using antibody raised against S. cerevisiae Idp1p detected two polypeptides consistent with these predictions. The functions of the predicted targetting sequences were confirmed by microscopic analysis of transformants containing fluorescent protein fusion constructs. Using anti-Idp1p antibodies, protein localization to mitochondria and peroxisomes was observed during growth on glucose while cytoplasmic and peroxisomal localization was found upon acetate or fatty acid induction. Therefore we have established that by the use of two transcription start points a single gene is sufficient to specify localization of NADP-dependent isocitrate dehydrogenase to three different cellular compartments in A. nidulans.
J. Biol. Chem, 10.1074/jbc.M105645200
Submitted on June 19, 2001
Revised on July 17, 2001
Accepted on July 31, 2001
A single gene produces mitochondrial, cytoplasmic and peroxisomal NADP-dependent isocitrate dehydrogenase in Aspergillus nidulans
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