Daucus carota cells contain specific DNA methyltransferase inhibitors that interfere with somatic embryogenesis

Results reported in this paper show that carrot cells contain a thermostable inhibiting activity for cytosine-5-DNA methyltransferase that, upon filtration chromatography, can be resolved into three major peaks. inhibiting activity was found in all plant species tested, though at a concentration lower than in carrot. These inhibiting activities differ in size, sensitivity to various hydrolytic treatments, specificity for DNA METases of eukaryotic and bacterial origin and kinetics of inhibition. Results of chemical analyses indicate that the inhibitors differ from lipidic inhibitors described in Escherichia coli and Streptomyces sp. and, given their sensitivity to proteinase K, appear to have a proteinaceous nature. The addition of these inhibitors (Sephaclex G25 peak II and peak III) to actively growing suspension rice cells reduced the rate of in vivo DNA methylation without interfering with DNA synthesis. Peak II also induced a general demethylation effect in carrot cell suspension, even if weaker than that caused by 5-azacytidine. Interestingly, inhibitors suppressed carrot embryogenesis but did not prevent undifferentiated cell proliferation of suspension cultures.