Vimentin's tail interacts with actin-containing structures in vivo.
The tail domain of the intermediate filament (IF) protein vimentin is unnecessary for IF assembly in vitro. To study the role of vimentin's tail in vivo, we constructed a plasmid that directs the synthesis of a 'myc-tagged' version of the Xenopus vimentin-1 tail domain in bacteria. This polypeptide, mycVimTail, was purified to near homogeneity and injected into cultured Xenopus A6 cells. In these cells the tail polypeptide co-localized with actin even in the presence of cytochalasin. Two myc-tagged control polypeptides argue for the specificity of this interaction. First, a similarly myc-tagged lamin tail domain localizes to the nucleus, indicating that the presence of the myc tag did not itself confer the ability to co-localize with actin (Hennekes and Nigg (1994) J. Cell Sci. 107, 1019-1029). Second, a myc-tagged polypeptide with a molecular mass and net charge at physiological pH (i.e. -4) similar to that of the mycVimTail polypeptide, failed to show any tendency to associate with actin-containing structures, indicating that the interaction between mycVimTail and actin-containing structures was not due to a simple ionic association. Franke (1987; Cell Biol. Int. Rep. 11, 831) noted a similarity in the primary sequence between the tail of the type I keratin DG81A and vimentin. To test whether the DG81A tail interacted with actin-containing structures, we constructed and purified myc-tagged DG81A tail polypeptides. Unexpectedly, these keratin tail polypeptides were largely insoluble under physiological conditions and formed aggregates at the site of injection. While this insolubility made it difficult to determine if they associated with actin-containing structures, it does provide direct evidence that the tails of vimentin and DG81A differ dramatically in their physical properties. Our data suggest that vimentin's tail domain has a highly extended structure, binds to actin-containing structures and may mediate the interaction between vimentin filaments and microfilaments involved in the control of vimentin filament organization (Hollenbeck et al. (1989) J. Cell Sci. 92, 621; Tint et al. (1991) J. Cell Sci. 98, 375).
University of Colorado, Boulder 80309-0347.
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Cary
M W
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Klymkowsky
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Evans
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A
Domingo
J A
JA
Dent
L E
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Backhus
D000199:Actins / Q000378:metabolism*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001665:Binding Sites; D002478:Cells, Cultured; D055598:Chemical Phenomena; D002627:Chemistry, Physical; D003571:Cytochalasin B / Q000494:pharmacology; D016259:Genes, myc; D008958:Models, Molecular; D008969:Molecular Sequence Data; D011485:Protein Binding; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D014746:Vimentin / Q000737:chemistry / Q000378:metabolism*; D014981:Xenopus
D000199:Actins; D011993:Recombinant Fusion Proteins; D014746:Vimentin; D003571:Cytochalasin B