A Cycloheximide-Sensitive Step in Transverse Microtubule Array Patterning

The growth properties of individual cells within a tissue determine plant morphology, and the organization of the cytoskeleton, particularly the microtubule arrays, determines cellular growth properties. We investigated the mechanisms governing the formation of transverse microtubule array patterns in axially growing Arabidopsis (Arabidopsis thaliana) epidermal hypocotyl cells. Using quantitative imaging approaches, we mapped the transition of the cortical microtubule arrays into a transverse coaligned pattern after induction with auxin and gibberellic acid. Hormone induction led to an early loss of microtubule plus end density and a rotation toward oblique patterns. Beginning 30 min after induction, transverse microtubules appeared at the cell's midzone concurrently with the loss of longitudinal polymers, eventually progressing apically and basally to remodel the array pattern. Based on the timing and known hormone-signaling pathways, we tested the hypothesis that the later events require de novo gene expression and, thus, constitute a level of genetic control over transverse patterning. We found that the presence of the translation inhibitor cycloheximide (CHX) resulted in a selective and reversible loss of transverse patterns that were replaced with radial-like pinwheel arrays exhibiting a split bipolar architecture centered at the cell's midzone. Experiments using hormone induction and CHX revealed that pinwheel arrays occur when transverse microtubules increase at the midzone but longitudinal microtubules in the split bipolar architecture are not suppressed. We propose that a key regulatory mechanism for creating the transverse microtubule coalignment in axially growing hypocotyls involves the expression of a CHX-sensitive factor that acts to suppress the nucleation of the longitudinally oriented polymers.