Identification of a distinct soluble subunit of an intermediate filament protein: tetrameric vimentin from living cells.
AUTOR(ES)
Soellner, P
RESUMO
Intermediate-sized filaments (IF) are among the most insoluble intracellular protein polymer structures. We have analyzed the small amounts of soluble vimentin, an IF protein, present in cytosol fractions obtained from lysis of cultured cells [rat RVF-SM cells, simian virus 40-transformed human fibroblasts, and human rhabdomyosarcoma (RD line) cells]. The molecular form of this soluble vimentin was determined by sucrose density gradient centrifugation, using vimentin-specific antibodies for subsequent ELISA and immunoblotting analyses. The majority of the soluble vimentin appeared in a distinct form indistinguishable in its sedimentation behavior from reconstituted tetrameric subunits of purified vimentin arrested at low ionic strength. The tetrameric coiled-coil nature of the soluble form of vimentin was indicated by the digestion pattern with chymotrypsin and by chemical crosslinking with copper-1,10-phenanthroline and dimethylsuberimidate. The competence of this soluble vimentin to assemble into IF at higher salt concentrations was demonstrated by electron microscopy. Pulse-chase experiments showed that the soluble form was not an exclusively posttranslational intermediate. We propose that in the living cell a small pool of a distinct soluble tetrameric form of vimentin exists which may exchange with polymeric IF vimentin.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=390883Documentos Relacionados
- Identification of glial filament protein and vimentin in the same intermediate filament system in human glioma cells.
- Alteration of vimentin intermediate filament expression during differentiation of human hemopoietic cells.
- Intermediate filament cDNAs from BHK-21 cells: demonstration of distinct genes for desmin and vimentin in all vertebrate classes.
- Subcellular localization of an intermediate filament protein and its mRNA in glial cells.
- Exoenzyme S of Pseudomonas aeruginosa ADP-ribosylates the intermediate filament protein vimentin.