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J. Cell Biol.,
Volume 143, Number 1, October 5, 1998 159-170

* Department of Cell and Molecular Biology, Northwestern University Medical School, Chicago, Illinois 60611; and The assembly and maintenance of an extended intermediate filament (IF) network in fibroblasts requires microtubule (MT) integrity. Using a
green fluorescent protein-vimentin construct, and
spreading BHK-21 cells as a model system to study
IF-MT interactions, we have discovered a novel mechanism involved in the assembly of the vimentin IF cytoskeleton. This entails the rapid, discontinuous, and
MT-dependent movement of IF precursors towards the
peripheral regions of the cytoplasm where they appear
to assemble into short fibrils. These precursors, or vimentin dots, move at speeds averaging 0.55 ± 0.24 µm/s. The vimentin dots colocalize with MT and their motility is inhibited after treatment with nocodazole.
Our studies further implicate a conventional kinesin
in the movement of the vimentin dots. The dots colocalize with conventional kinesin as shown by indirect immunofluorescence, and IF preparations from spreading
cells are enriched in kinesin. Furthermore, microinjection of kinesin antibodies into spreading cells prevents
the assembly of an extended IF network. These studies
provide insights into the interactions between the IF
and MT systems. They also suggest a role for conventional kinesin in the distribution of non-membranous protein cargo, and the local regulation of IF assembly.
Howard
Hughes Medical Institute, University of California, San Francisco, California 94143
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