MOLECULAR AND BIOCHEMICAL-CHARACTERIZATION OF HERBICIDE-RESISTANT MUTANTS OF CYANOBACTERIA REVEALS THAT PHYTOENE DESATURATION IS A RATE-LIMITING STEP IN CAROTENOID BIOSYNTHESIS

Mutant strains of the cyanobacterium Synechococcus sp. PCC 7942 that a re resistant to the herbicides norflurazon and fluridone were analyzed . These herbicides inhibit phytoene desaturase, a key enzyme in the ca rotenoid biosynthetic pathway. In three mutants the phenotype was rela ted to specific point mutations in pds, the gene encoding phytoene des aturase. Since the resistance was manifested in a cell-free carotenoge nic assay, it is evident that the predicted amino acid changes in the target protein alter the enzyme-herbicide interactions. In addition, t he mutations also reduced the in vitro activity of phytoene desaturase . Increased levels of the substrate phytoene, accompanied by a reducti on in colored carotenoids, were detected in cells of each of the mutan t strains. A correlation was established between the degree of increas e in the steady-state levels of phytoene and the extent of reduction i n total carotenoid content in the cells. These two phenomena in turn a re correlated with the rate of enzymatic activity of phytoene desatura se that was measured in vitro. Hence we deduce that phytoene desaturat ion is a rate-limiting step in carotenogenesis in cyanobacteria. Suppo rt for this conclusion is obtained from analysis of cells of an additi onal mutant strain, which overexpress phytoene desaturase due to a del etion mutation in the promoter region of pds. Cells of this mutant con tained more colored carotenoids than the wild-type and were resistant to herbicides that inhibit phytoene desaturase.