Eur J Cell Biol 1995 Feb 01;662:136-50.

Truncation mutagenesis of the non-alpha-helical carboxyterminal tail domain of vimentin reveals contributions to cellular localization but not to filament assembly.

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We have investigated the effect of stepwise truncating the carboxyterminal domain ("tail") of the intermediate filament (IF) protein vimentin of the clawed toad, Xenopus laevis, on filament assembly in vitro and, using cell transfection, in vivo and also on the cellular topology of the structures formed. All truncations examined, except the minimal one missing the last 11 amino acids which made the protein more sensitive to changes of ionic strength, did not significantly alter IF assembly in vitro, as judged by electron microscopy, viscometry and determination of viscoelastic properties with a laser-operated torsion pendulum. Stable transfections of vimentin-free mammalian cells with cDNAs encoding these mutations resulted at 28 degrees C, i.e. the permissive temperature for assembly of Xenopus vimentin, in the formation of extended IF bundle arrays. At 37 degrees C, however, the mutants lacking more than the last 35 amino acids could leave the cytoplasm and accumulated in the nucleus, indicating a certain topogenic element is located in the tail and directs cytoplasmic restriction in the wild-type protein although this does not form IFs under these conditions. Transfer to the nucleus is, however, abolished if the IF-consensus motif at the end of the rod domain is removed, suggesting that this part of the molecule also contributes to nuclear location. Similar results were obtained with human vimentin: While the rod entered the nucleus, headless vimentin, unable to form IFs, remained restricted to the cytoplasm owing to its tail domain. In contrast, tailless human vimentin and tailless mouse desmin, which are fully assembly-competent in vitro, both formed extensive IF arrays in the cytoplasm but did not accumulate in the nucleus. We conclude that in class III IF proteins stepwise deletions in the tail, while not considerably altering IF assembly in vitro, can change the topogenesis of IF proteins and structures in the living cell.

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Species referenced: Xenopus laevis
Genes referenced: nr2e1 vim