MOLECULAR-CLONING AND EXPRESSION OF CHLOROPLAST NADP-MALATE DEHYDROGENASE DURING CRASSULACEAN ACID METABOLISM INDUCTION BY SALT STRESS

A full-length cDNA clone for NADP+-dependent malate dehydrogenase (NADP-MDH; EC 1.1.1.82) from the facultative CAM plant, Mesembryanthemum crystallinum has been isolated and characterized. NADP-MDH is responsible for the reduction of oxaloacetate to malate in the chloroplasts of higher plants. The cDNA clone is 1747 bp in size and contains a single open reading frame encoding a 441 amino acid long precursor polypeptide with a predicted molecular weight of 47 949. The predicted, mature NADP-MDH polypeptide sequence from M. crystallinum shares 82.7% to 84% amino acid identity with other known higher plant sequences. Genomic Southern blot analysis of M. crystallinum DNA indicates that MDH is encoded by a small gene family. Steady-state transcript levels for chloroplast NADP-MDH decrease transiently in the leaves after salt stress and then increase to levels greater than two-fold higher than in unstressed plants. Transcript levels in roots are extremely low and are unaffected by salt-stress treatment. In vitro transcription run-on experiments using isolated nuclei from leaf tissue confirm that the accumulation of NADP-MDH transcripts is, at least in part, the result of increased transcription of this gene during salt stress. The salt-stress-induced expression pattern of this enzyme suggests that it may participate in the CO2 fixation pathway during CAM.