Molecular cloning of glutamate dehydrogenase genes of Nicotiana plumbaginifolia: structure analysis and regulation of their expresion by physiological and stress conditions

Two Nicotiana plumbaginifolia gdh genes (gdhA and gdhB) encoding for NADH dependent glutamate dehydrogenase (GDH) were isolated by PCR and I-PCR (inverse-PCR) and sequenced. The coding sequence is interrupted by eight introns that are located in the same positions in both genes. Computer analysis of the promoter regions of gdhA (650 nt) and gdhB (1800 nt) revealed several conserved motifs and, for gdhB, the presence of an I-Box element known to have light dependent regulatory functions in plants. The response of the gdh genes to carbohydrate deprivation and to ammonium feeding and to various stress conditions (salinity, temperature shifts and heavy metals) was examined in callus culture. The results clearly showed that the carbohydrate status of the cell is the main factor affecting gdh genes expression. Stress condition may depress or increase gdh transcript levels, but theses are not always correlated to the corresponding levels of GDH activity. Altogether these results point to the existence of multiple levels of regulation (transcriptional and post-transcriptional) of gdh genes.