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J. Cell Biol.,
Volume 140, Number 2, January 26, 1998 391-401
* Department of Anatomy and Program in Cellular and Molecular Biology, The University of Wisconsin Medical School,
Madison, Wisconsin 53706; and Previous work from our laboratory suggested that microtubules are released from the neuronal centrosome and then transported into the axon
(Ahmad, F.J., and P.W. Baas. 1995. J. Cell Sci. 108:
2761-2769). In these studies, cultured sympathetic neurons were treated with nocodazole to depolymerize
most of their microtubule polymer, rinsed free of the
drug for a few minutes to permit a burst of microtubule
assembly from the centrosome, and then exposed to nanomolar levels of vinblastine to suppress further microtubule assembly from occurring. Over time, the microtubules appeared first near the centrosome, then
dispersed throughout the cytoplasm, and finally concentrated beneath the periphery of the cell body and
within developing axons. In the present study, we microinjected fluorescent tubulin into the neurons at the
time of the vinblastine treatment. Fluorescent tubulin
was not detected in the microtubules over the time
frame of the experiment, confirming that the redistribution of microtubules observed with the experimental
regime reflects microtubule transport rather than microtubule assembly. To determine whether cytoplasmic
dynein is the motor protein that drives this transport,
we experimentally increased the levels of the dynamitin subunit of dynactin within the neurons. Dynactin, a
complex of proteins that mediates the interaction of cytoplasmic dynein and its cargo, dissociates under these
conditions, resulting in a cessation of all functions of
the motor tested to date (Echeverri, C.J., B.M. Paschal,
K.T. Vaughan, and R.B. Vallee. 1996. J. Cell Biol. 132:
617-633). In the presence of excess dynamitin, the microtubules did not show the outward progression but
instead remained near the centrosome or dispersed
throughout the cytoplasm. On the basis of these results,
we conclude that cytoplasmic dynein and dynactin are
essential for the transport of microtubules from the
centrosome into the axon.
Cell Biology Group, Worcester Foundation for Biomedical Research, Shrewsbury,
Massachusetts 01545
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