A branched-chain amino acid aminotransferase gene isolated from Hordeum vulgare is differentially regulated by drought stress
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Differential display was used to isolate cDNA clones showing differential expression in response to ABA, drought and cold in barley seedling shoots. One drought-regulated cDNA clone (DD12) was further analyzed and found to encode a branched-chain amino acid aminotransferase (HvBCAT-1). A genomic clone was isolated by probing the Morex BAC library with the cDNA clone DD12 and the structure of Hvbcat-1 was elucidated. The coding region is interrupted by six introns and contains a predicted mitochondrial transit peptide. Hvbcat1 was mapped to chromosome 4H. A comparison was made to rice and Arabidopsis genes to identify conserved structural patterns. Complementation of a yeast (Saccharomyces cerevisiae) double knockout strain revealed that HvBCAT-1 can function as the mitochondrial (catabolic) BCATs in vivo. Transcript levels of Hvbcat-1, increased in response to drought stress. As the first enzyme in the branched-chain amino acid (BCAA) catabolic pathway, HvBCAT-1 might have a role in the degradation of BCAA. Degradation of BCAA could serve as a detoxification mechanism that maintains the pool of free branched-chain amino acids at low and non toxic levels, under drought stress conditions.
We are grateful to prof. R. Lill and U. Muehlenhoff (University of Munchen, Germany) for the generous gift of the Δbat2/gal-bat1 yeast strain and to prof. Tiziana Lodi (University of Parma, Italy) for the gift of the plasmid pYeDP10 for yeast transformation. This work has been supported by NATO Grant (CLG 978261) to N. Marmiroli, by project “Biotecnologie Vegetali” (MIPA) to N. Marmiroli, by CNR-Agenzia 2000 project to M. Gulli and in part by NSF DBI-0321756, “Coupling Expressed Sequences and Bacterial Artificial Chromosome Resources to Access the Barley Genome” to T.J. Close.
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