INCREASED CELL CYCLE-DEPENDENT STAINING OF PLANT-CELLS BY THE ANTIBODY MPM-2 CORRELATES WITH PREPROPHASE BAND FORMATION

Cytoplasmic microtubules of animal cells catastrophically depolymerize upon entry into mitosis but in higher plants there is a longer transi tion during which cortical microtubules form an increasingly narrow pr eprophase band, and the chromatin gradually condenses. Progression tow ards mitosis in onion root tip cells was analysed using a CCD camera a nd image processing to quantify fluorescence staining by the monoclona l antibody MPM-2, which recognizes mitotic phosphoproteins in a range of eukaryotic cells. MPM-2 fluorescence, which was predominantly nucle ar, was categorized relative to the stage of the DNA cycle (using DAPI ), and to the microtubule cycle (using anti-tubulin) in individual cel ls. Cells with the characteristic interphase cortical microtubule arra ys had a bimodal distribution of DAPI fluorescence, indicating that so me were in G1 (2C DNA) whilst the double value suggested the others to be in G2 (4C). There was no difference in MPM-2 fluorescence between 2C and 4C cells possessing the cortical array in which microtubules we re evenly distributed. However, in 4C cells possessing a preprophase b and MPM-2 values doubled; this relationship applied not only to tight PPBs but to early, broad PPBs in which the individual microtubules cou ld still be distinguished. Since alkaline phosphatase abolished MPM-2 reactivity it is concluded that mitotic phosphoproteins do not necessa rily begin to accumulate in G2 per se, but during that part of G2 when the preprophase band first becomes recognizable as a distinct entity.