RanBP10 is dispensable for spontaneous microtubule polymerization but is essential for marginal band contraction after platelet activation. (A) Microtubule depolymerization and repolymerization was normal after platelet cooling and rewarming. Tubulin is stained in red (Alexa647) and Ran in green (Alexa488). (B) TEM ultragraphs of platelets treated with 1mM or 0.1mM PAR4p show that in wild-type platelets the microtubule coil contracts (arrows, i and ii) and granules become centralized. In RanBP10^−/− platelets (iii and iv) the marginal microtubule band fails to contract and granules are no longer centralized (arrowheads). Normal platelet activation is shown by protrusions in either strain. Subpanel iv shows an example of disorganized microtubule filaments (arrow). (C) Immunofluorescence staining with anti–β1-tubulin antibody and quantification of fixed platelets at time-points indicated after stimulation with PAR4p (top panel). Quantification of wild-type (black) and RanBP10^−/− platelets (gray) after stimulation with 0.1mM PARp (hatched bars) and 1mM PAR4p (filled bars; ***P < .001, double-sided Student t test with independent variances).