Effects of DNA on Microtubule Assembly

A preliminary report on parts of this work was presented at the 12th FEBS Meeting (Dresden), July 2-8, 1978. The effects of microtubule-associated protein binding to DNA on microtubule assembly in vitro were studied by turbidity measurements and electron microscopy. DNA inhibited microtubule assembly in a concentration-dependent manner. At DNA concentrations low enough to allow some polymerisation to occur, assembled microtubules were longer than those formed in the absence, indicating a reduction in the number of microtubule-associated proteins available for polymerisation. Also, by binding microtubule-associated proteins, DNA raised the critical concentration of microtubule protein necessary for microtubule assembly. However, satellite DNA, which bound microtubule-associated proteins more effectively than bulk DNA, raised the critical concentration less. Similar effects were observed for the synthetic DNA polymers poly(dA) poly(dT) and poly(dA-dT). poly(dA-dT) compared to poly(dG). poly(dC) and poly(dG-dC). poly(dG-dC). Nitrocellulose filter binding assays showed that only microtubule-associated protein fractions which contained high-molecular-weight components exhibited DNA binding specifity. These high-molecular-weight polypeptides bound preferentially to eukariotic DNA as compared to phage DNA; fractions containing tau protein bound equally, well to both DNA species. High-molecular-weight polypeptides also showed preferential binding to poly(dA-dT). poly(dA-dT) compared to poly(dG-dC). poly(dG-dC) synthetic DNA polymers. We suggest that satellite DNA acts in vivo as an initiation site for microtubule formation and that high-molecular-weight polypeptides could be the linker components of the kinetochore. This work has been carried out with the support of grants from the Comision Asesora para el Desarollo de la Investigacion Cientifica y Tecnica, the Comision Administradora del Descuento Complementario (I.N.P.) and the Austrian Fonds zur Forderung der wissenschaftlichen Forschung.