A rat monoclonal antibody reacting specifically with the tyrosylated form of alpha-tubulin. I. Biochemical characterization, effects on microtubule polymerization in vitro, and microtubule polymerization and organization in vivo.
The antigenic site recognized by a rat monoclonal antibody (clone YL 1/2) reacting with alpha-tubulin (Kilmartin, J.V., B. Wright, and C. Milstein, 1982, J. Cell Biol., 93:576-582) has been determined and partially characterized. YL 1/2 reacts specifically with the tyrosylated form of brain alpha-tubulin from different mammalian species. YL 1/2 reacts with the synthetic peptide Gly-(Glu)3-Gly-(Glu)2-Tyr, corresponding to the carboxyterminal amino acid sequence of tyrosylated alpha-tubulin, but does not react with Gly-(Glu)3-Gly-(Glu)2, the constituent peptide of detyrosylated alpha-tubulin. Electron microscopy as well as direct and indirect immunofluorescence microscopy shows that YL 1/2 binds to the surface of microtubules polymerized in vitro and in vivo. Further in vitro studies show that the antibody has no effect on the rate and extent of microtubule polymerization, the stability of microtubules, and the incorporation of the microtubule-associated proteins (MAP2) and tau into microtubules. In vivo studies using Swiss 3T3 fibroblasts injected with YL 1/2 show that; when injected at low concentration (2 mg IgG/ml in the injection solution), the antibody binds to microtubules without changing their distribution in the cytoplasm. Injection of larger concentration of YL 1/2 (6 mg IgG/ml) induces the formation of microtubule bundles, and still higher concentrations cause the aggregation of microtubule bundles around the nucleus (greater than 12 mg IgG/ml).
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D005347:Fibroblasts / Q000648:ultrastructure; D005455:Fluorescent Antibody Technique; D051379:Mice; D008854:Microscopy, Electron; D008869:Microtubule-Associated Proteins; D008870:Microtubules / Q000378:metabolism* / Q000648:ultrastructure; D011108:Polymers / Q000378:metabolism; D011506:Proteins / Q000378:metabolism; D051381:Rats; D013552:Swine; D014404:Tubulin / Q000031:analogs & derivatives* / Q000276:immunology; D016875:tau Proteins
D000911:Antibodies, Monoclonal; D008869:Microtubule-Associated Proteins; D011108:Polymers; D011506:Proteins; D014404:Tubulin; D016875:tau Proteins; C039520:tyrosine-tubulin