CHARACTERIZATION OF VITIS-VINIFERA L GLUTAMINE-SYNTHETASE AND MOLECULAR-CLONING OF CDNAS FOR THE CYTOSOLIC ENZYME

Grapevine (Vitis vinifera L.) glutamine synthetase (GS) was analysed i nto two distinct classes of isoforms; one of them was present in both leaf and root tissues while the other one showed leaf specificity. Wes tern blot analysis revealed that grapevine GS consists of three types of polypeptides of distinct size and differential tissue specificity. Two structurally distinct cDNA clones, pGS1;1 and pGS1;2, encoding gra pevine GS were isolated from a cell suspension library and characteriz ed. Both clones contained open reading frames encoding for polypeptide s of 356 amino acids with a predicted molecular mass of about 39 kDa. Although the coding sequences of pGS1;1 and pGS1;2 were 84% similar, t heir 5'- and 3'-untranslated sequences showed only 40% similarity. The coding sequences of the two clones and the derived amino acid sequenc es showed higher homology to cytosolic than to chloroplastic GSs of ot her higher plants indicating that the cDNAs isolated encode for cytoso lic isoforms of grapevine GS. Southern blot analysis suggested the exi stence of more than two GS genes in the grapevine genome. In northern blots both clones were hybridized to mRNAs of about 1.4 kb that are di fferentially expressed in the various tissues. Supply of nitrate or am monium in the cell suspension culture medium, as a sole nitrogen sourc e,resulted in differential response of the pGS1;1- and pGS1;2-related genes.