Reconstitution of intermediate filaments from a higher plant.
Immunological studies have shown that plants contain intermediate-filament antigens, but it is not known whether these proteins are capable in themselves of forming filaments. To address this problem, a detergent-resistant and high-salt-insoluble fraction from carrot (Daucus carota L.) suspension cells was solubilized with 9 M-urea and then subjected to a two-step dialysis procedure, devised for the reconstitution of animal intermediate filaments. This induced the self-assembly of 10 nm filaments and large bundles of filaments. The predominant components of reconstituted material were polypeptides with apparent molecular masses between 58 and 62 kDa. These polypeptides immunoblotted with two monoclonal antibodies known to show broad cross-reactivity with intermediate filaments across the phylogenetic spectrum. This establishes that the antigens are able to self-assemble into intermediate-sized filaments.
Department of Cell Biology, John Innes Institute, Norwich, U.K.
A J
AJ
Hargreaves
K C
KC
Goodbody
C W
CW
Lloyd
D003599:Cytoskeleton / Q000648:ultrastructure*; D004591:Electrophoresis, Polyacrylamide Gel; D015151:Immunoblotting; D007382:Intermediate Filaments / Q000648:ultrastructure*; D008854:Microscopy, Electron; D014674:Plant Proteins, Dietary / Q000378:metabolism*
D014674:Plant Proteins, Dietary