COLOCALIZATION OF CYTOKININS WITH PROTEINS RELATED TO CELL-PROLIFERATION IN DEVELOPING SOMATIC EMBRYOS OF DACTYLIS-GLOMERATA L

The present report provides evidence for co-localization of cytokinins with cell proliferation-associated nuclear proteins. Somatic embryos of Dactylis glomerata in two stages of development are used as a model system comprising both proliferating and initially differentiated cel ls. Cytokinins are localized using antibodies with marked specificity against isopentenyladenine/adenosine (2iP/2iPA) or zeatin/ riboside (Z /ZR). The proliferation-associated nuclear antigen, mitotin, is analys ed using a specific monoclonal antibody. The nuclear protein BM28, req uired for the onset of DNA replication and for cell division is identi fied by an affinity-purified polyclonal antibody. Using double immunof luorescence labelling with the antibodies against cytokinins and again st each of the nuclear proteins, immunoreaction is observed generally in the same nuclei of almost all cells in globular embryos and in the nuclei of cells in meristematic areas of the more developed embryos. O nly small numbers of individual nuclei positive for both type of antib odies were found in the surrounding vacuolated parenchymatous cells. T he occurrence of plant antigens homologous to BM28 and mitotin is conf irmed by immunoblotting assay. In SDS-PAGE blots the anti-BM28 antibod y reacts with a protein of 58 kDa. The anti-mitotin antibody recognize s several (160, 140, 125, 93, and 80 kDa) polypeptides. The data showi ng nuclear co-localization of cytokinins and proteins with a suggested role in the onset of DNA synthesis and in cell division provide a new base for further studyon the mode of action of cytokinins in cell cyc le regulation.