Daucus carota Cell Cultures: Polyamines and Effect of Polyamine Biosynthesis Inhibitors in the Preembryogenic Phase and Different Embryo Stages

Summary Polyamine content, ornithine decarboxylase (EC 4.1.1.17) and arginine decarboxylase (EC 4.1.1.19) activities, as well as the effects of two inhibitors of putrescine synthesis, were studied during the preembryogenic phase and different stages of somatic embryogenesis in suspension cultures carrots ( Daucus carota L.). In the preembryogenic phase α-difluoromethylornithine at 5mM, an irreversible inhibitor of ornithine decarboxylase, does not reduce growth but causes inhibition of putrescine accumulation; canavanine, a competitive inhibitor of arginine decarboxylase, completely blocks growth in the same phase. Difluoromethylornithine seems to prolong the commitment of the cells to embryogenesis during the preembryogenic phase, while canavanine inhibits the subsequent embryo differentiation. In general, a decrease in polyamine content and higher ornithine decarboxylase as compared to arginine decarboxylase activity is observed during the preembryogenic phase. Difluoromethylornithine does not affect ornithine decarboxylase in vivo , while both difluoromethylornithine and canavanine inhibit ornithine and arginine decarboxylase, respectively, in vitro . Spermidine and spermine contents, expressed on a per embryo basis, increase during the progression from globular to heart and torpedo stages of embryogenesis. While putrescine is the main polyamine in the heart stage, spermidine is more abundant in the torpedo stage. Embryos treated with difluoromethylornithine for 36h develop into normal plantlets at a higher frequency, and they are consistently larger than controls; the canavanine treatment blocks any further development of the embryos, which are always abnormal.