Colchicine binding in the free-living nematode Caenorhabditis elegans

Abstract

The [ 3H]colchicine-binding activity of a crude supernatant of the free-living nematode Caenorhabditis elegans was resolved into a non-saturable component and a tubulin-specific component after partial purification of tubulin by polylysine affinity chromatography. The two fractions displayed opposing thermal dependencies of [ 3H]colchicine binding, with non-saturable binding increasing, and tubulin binding decreasing, at 4°C. Binding of [ 3H]colchicine to C. elegans tubulin at 37°C is a pseudo-first-order rate process with a long equilibration time. The affinity of C. elegans tubulin for [ 3H]colchicine is relatively low ( K a = 1.7 · 10 5 M −1) and is characteristic of the colchicine binding affinities observed for tubulins derived from parasitic nematodes. [ 3H]Colchicine binding to C. elegans tubulin was inhibited by unlabelled colchicine, podophyllotoxin and mebendazole, and was enhanced by vinblastine. The inhibition of [ 3H]colchicine binding by mebendazole was 10-fold greater for C. elegans tubulin than for ovine brain tubulin. The inhibition of [ 3H]colchicine binding to C. elegans tubulin by mebendazole is consistent with the recognised anthelmintic action of the benzimidazole carbamates. These data indicate that C. elegans is a useful model for examining the interactions between microtubule inhibitors and the colchicine binding site of nematode tubulin.