Microtubule polymer assembly and transport during axonal elongation.
As axons elongate, tubulin, which is synthesized in the cell body, must be transported and assembled into new structures in the axon. The mechanism of transport and the location of assembly are presently unknown. We report here on the use of tubulin tagged with a photoactivatable fluorescent group to investigate these issues. Photoactivatable tubulin, microinjected into frog embryos at the two-cell stage, is incorporated into microtubules in neurons obtained from explants of the neural tube. When activated by light, a fluorescent mark is made on the microtubules in the axon, and transport and turnover can be visualized directly. We find that microtubules are generated in or near the cell body and continually transported distally as a coherent phase of polymer during axon elongation. This vectorial polymer movement was observed at all levels on the axon, even in the absence of axonal elongation. Measurements of the rate of polymer translocation at various places in the axon suggest that new polymer is formed by intercalary assembly along the axon and assembly at the growth cone in addition to transport of polymer from the cell body. Finally, polymer movement near the growth cone appeared to respond in a characteristic manner to growth cone behavior, while polymer proximally in the axon moved more consistently. These results suggest that microtubule translocation is the principal means of tubulin transport and that translocation plays an important role in generating new axon structure at the growth cone.
Reinsch
S S
SS
Department of Biochemistry and Biophysics, University of California, San Francisco 94143.
Mitchison
T J
TJ
Kirschner
M
M
D000818:Animals; D001369:Axons / Q000378:metabolism* / Q000648:ultrastructure; D001692:Biological Transport; D003911:Dextrans / Q000378:metabolism; D019793:Fluorescein; D005452:Fluoresceins / Q000378:metabolism; D007700:Kinetics; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000378:metabolism* / Q000648:ultrastructure; D009474:Neurons / Q000166:cytology*; D014404:Tubulin / Q000378:metabolism; D014981:Xenopus / Q000196:embryology
D003911:Dextrans; D005452:Fluoresceins; D014404:Tubulin; D019793:Fluorescein