Alteration of Microtubule Dynamic Instability during Preprophase Band Formation Revealed by Yellow Fluorescent Protein–CLIP170 Microtubule Plus-End Labeling[W]

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Abstract
At the onset of mitosis, plant cells form a microtubular preprophase band that defines the plane of cell division, but the mechanism of its formation remains a mystery. Here, we describe the use of mammalian yellow fluorescent protein-tagged CLIP170 to visualize the dynamic plus ends of plant microtubules in transfected cowpea protoplasts and in stably transformed and dividing tobacco Bright Yellow 2 cells. Using plus-end labeling, we observed dynamic instability in different microtubular conformations in live plant cells. The interphase plant microtubules grow at 5 micro m/min, shrink at 20 micro m/min, and display catastrophe and rescue frequencies of 0.02 and 0.08 events/s, respectively, exhibiting faster turnover than their mammalian counterparts. Strikingly, during preprophase band formation, the growth rate and catastrophe frequency of plant microtubules double, whereas the shrinkage rate and rescue frequency remain unchanged, making microtubules shorter and more dynamic. Using these novel insights and four-dimensional time-lapse imaging data, we propose a model that can explain the mechanism by which changes in microtubule dynamic instability drive the dramatic rearrangements of microtubules during preprophase band and spindle formation in plant cells.