Comparison of spindle elongation in vivo and in vitro in Stephanopyxis turris

Abstract

The spindle in dividing cells of the diatom Stephanopyxis turris contains three distinct classes of microtubules: central spindle microtubules, which slide over each other and grow during anaphase spindle elongation; kinetochore-attached microtubules, which are located on the outer surface of the central spindle; and peripheral microtubules, which fan out from the spindle poles in astral-like arrays. The poles are multilayered structures, which remain attached to the spindle after isolation. In vitro, after addition of ATP, central spindles elongate and the two half-spindles slide completely apart with a concurrent decrease in the extent and magnitude of the zone of microtubule overlap. Spindle elongation takes place in spindles whose chromatin has been removed by enzymic digestion and the extent of elongation in vitro is increased by the addition of neurotubulin. After ATP addition the arrays of interdigitating microtubules in the zone of overlap become disordered and selectively depolymerize from the overlap zone polewards. In some reactivated spindles an unusual structure, a striated fibre, can be seen running from the pole plates part of the way towards the spindle midzone. The fibre has no precedent in mitotic ultrastructure and its function is unclear. These results demonstrate that we can duplicate the essential elements of anaphase B in vitro and that this system will be useful for further studies of the molecular basis of spindle elongation.