Strongylocentrotus purpuratus Spindle Tubulin. II. Characteristics of Its Sensitivity to Ca++ and the Effects of Calmodulin Isolated from Bovine Brain and S. purpuratus Eggs

• Published in: The Journal of cell biology 1982-06, Vol.93 (3), p.797-803
• Main Authors: Thomas C. S. Keller, III, Jemiolo, David K., Burgess, Wilson H., Rebhun, Lionel I.
• Format: Article
• Language: English
• Subjects: Animals; Biopolymers; Brain Chemistry; Calcium - metabolism; Calcium-Binding Proteins - pharmacology; Calmodulin - pharmacology; Cattle; Depolymerization; Eggs; Embryos; In Vitro Techniques; Microtubules; Mitosis; Mitotic spindle apparatus; Muscle spindles; Physiological regulation; Polymerization; Polymers; Sea Urchins - metabolism; Temperature; Tubulin - isolation & purification; Tubulin - metabolism; Ungulates
• ISSN: 0021-9525; 1540-8140
• DOI: 10.1083/jcb.93.3.797

Tubulin was extracted from spindles isolated from embryos of the sea urchin Strongylocentrotus purpuratus and purified through cycles of temperature-dependent assembly and disassembly. At 37°C, the majority of the cycle-purified spindle tubulin polymer is insensitive to free Ca++ at concentrations below 0.4 mM, requiring free Ca++ concentrations >1 mM for complete depolymerization. However, free Ca++ at concentrations above 1 μM inhibits initiation of polymer formation without significantly inhibiting the rate of elongation onto existing polymer. At 15°C and 18°C, temperatures that are physiological for S. purpuratus embryos, spindle tubulin polymer is sensitive to free Ca++ at micromolar concentrations such that 3-20 μM free Ca++ causes complete depolymerization. Calmodulin purified from either bovine brain or S. purpuratus eggs does not affect the Ca++ sensitivity of the spindle tubulin at 37°C, although both increase the Ca++ sensitivity of cycle-purified bovine brain tubulin. These results indicate that cycle-purified spindle tubulin and cycle-purified bovine brain tubulin differ significantly in their responses to calmodulin and in their Ca++ sensitivities at their physiological temperatures. They also suggest that, in vivo, spindle tubulin may be regulated by physiological levels of intracellular Ca++ in the absence of Ca++-sensitizing factors.