a1A and a1C form microtubules to display distinct properties mainly mediated by their C-terminal tails

Abstract

Microtubules consisting of a/b-tubulin dimers play critical roles in cells. More than seven genes encode a-tubulin in vertebrates. However, the property of microtubules composed of different a-tubulin isotypes is largely unknown. Here, we purified recombinant tubulin heterodimers of mouse a-tubulin isotypes including a1A and a1C with b-tubulin isotype b2A. In vitro microtubule reconstitution assay detected that a1C/b2A microtubules grew faster and underwent catastrophe less frequently than a1A/b2A microtubules. Generation of chimeric tail-swapped and point-mutation tubulins revealed that the carboxyl-terminal (C-terminal) tails of a-tubulin isotypes largely accounted for the differences in polymerization dynamics of a1A/b2A and a1C/b2A microtubules. Kinetics analysis showed that in comparison to a1A/b2A microtubules, a1C/b2A microtubules displayed higher on-rate, lower off-rate, and similar GTP hydrolysis rate at the plus-end, suggesting a contribution of higher plus-end affinity to faster growth and less frequent catastrophe of a1C/b2A microtubules. Furthermore, EB1 had a higher binding ability to a1C/b2A microtubules than to a1A/b2A ones, which could also be attributed to the difference in the C-terminal tails of these two a-tubulin isotypes. Thus, a-tubulin isotypes diversify microtubule properties, which, to a great extent, could be accounted by their C-terminal tails.