Okadaic acid blocks the cell cycle at early mitosis in suspension cultures ofNicotiana plumbaginifolia. Nuclear DNA content was measured in treated cells by propidium iodide staining, fluorescence microscopy and quantitative analysis of the video image. Nuclear DNA levels in inhibited populations showed that cells continued to progress from G1 phase through S phase and accumulated in G2 phase. Arrested cells in 12μM okadaic acid had a condensed chromatin configuration and persisting nucleolus similar to normal early prophase. Progress to early prophase was also indicated by development of the preprophase band (PPB) of microtubules. PPB microtubules disassembled in 95%of the inhibited cells with the same timing as in control cells, although the treated cells did not progress to prometaphase mitotic spindle assembly that normally precedes PPB breakdown, therefore okadaic acid can disrupt the normal dependence of PPB disassembly on prometaphase nuclear events and indicates that the normal signal for disassembly may be an increase in phosphorylation of PPB associated proteins. Okadaic acid at 12μM caused increased levels of phosphorylated proteins, in particular those of 108 kDa, 49 kDa, 36 kDa, 33 kDa, 31 kDa, but more complex effects on some phosphoproteins were indicated by reductions in a phosphoprotein of 41 kDa and one of approximately 190 kDa. It is concluded that the mitotic phase of the plant cell cycle is more sensitive than preceding cycle phases to the disruption of protein phosphorylation levels by okadaic acid and it is proposed that the inhibitor blocks division by interfering with essential changes in the phosphorylation state of proteins at mitosis. This conclusion is discussed in relation to genetical and biochemical evidence that protein kinases and phosphatases are involved in the cell division of plants and other eukaryote
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