In the early 1960s, microtubules were known to be constituents of the mitotic spindle fibres (see Milestone 5) and the 9+2 array of filaments that are observed in cilia and spermatozoa tails (see Milestone 4). The identification of tubulin as the basic subunit of microtubules opened up these structures to molecular analysis and demonstrated that microtubules from different sources had the same composition. The drug colchicine played a key role in this discovery.
Today, colchicine, together with colcemid and nocodazole, is commonly used in the laboratory to block microtubule polymerization; these drugs bind to tubulin and prevent its addition to growing microtubule ends. In the 1960s, although colchinine was known to destroy the mitotic spindle, a confusing body of literature described other cellular and physiological effects.
The identification of tubulin as the basic subunit of microtubules opened up these structures to molecular analysis...
In 1967, Edward Taylor reported that the kinetics of colchicine binding to cells could be modelled by a single class of binding sites, indicating that a unique target might exist. Gary Borisy embarked on the project to identify it. By adding radiolabelled colchicine to a range of extracts from cells and tissues, he found a single 6S component co-purifying with colchicine. Importantly, this binding activity was high in tissue-culture cells, sea urchin eggs, isolated mitotic spindles and brain tissue — all of which are rich in microtubules. Borisy and Taylor therefore proposed that the 6S protein was the microtubule subunit, although the name tubulin was coined only in a later report by Hideo Morhi on the biochemical composition of spermatozoa flagella. In addition to the discovery of tubulin, the work by Borisy and Taylor established the powerful approach of using specific drugs to probe the function of the cytoskeleton.
Efforts to isolate tubulin and to study its assembly properties ensued. In 1972, Richard Weisenberg and Borisy re-assembled microtubules from tubulin purified from rat brain homogenates, showing that the process required a calcium chelator. This key study demonstrated that the capacity for assembly resided in the tubulin subunit itself and did not require a separate polymerase. Work by Weisenberg also provided a protocol for microtubule self-assembly, opening the door to research into its mechanisms (see Milestone 14).
