Isolation of a 90-kD microtubule-associated protein from tobacco membranes

The organization and function of microtubules in plant cells are important in key developmental events, including the regulation of directional cellulose deposition. Bridges connecting microtubules to each other and to membranes and other organelles have been documented by electron microscopy; howev...

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Bibliographic Details
Published in:The Plant cell 1996-11, Vol.8 (11), p.2127-2138
Main Authors: Marc, J. (University of Sydney, Sydney, Australia.), Sharkey, D.E, Durso, N.A, Zhang, M, Cyr, R.J
Format: Article
Language:English
Subjects:
Antibodies
BIOCHIMIE
BIOQUIMICA
Carrier proteins
CITOPLASMA
CYTOPLASME
DAUCUS CAROTA
ESTRUCTURA CELULAR
Gels
GLICOPROTEINAS
GLYCOPROTEINE
HeLa cells
IMMUNOLOGIE
INMUNOLOGIA
MEMBRANAS CELULARES
MEMBRANE CELLULAIRE
Microscopy
Microtubule associated proteins
Microtubules
Monoclonal antibodies
NICOTIANA TABACUM
Plant cells
PROTEINAS AGLUTINANTES
PROTEINE DE LIAISON
Protoplasts
STRUCTURE CELLULAIRE
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Summary:The organization and function of microtubules in plant cells are important in key developmental events, including the regulation of directional cellulose deposition. Bridges connecting microtubules to each other and to membranes and other organelles have been documented by electron microscopy; however, the biochemical and molecular nature of these linkages is not known. We have partitioned proteins from a suspension culture of tobacco into cytosolic and membrane fractions, solubilized the membrane fraction with a zwitterionic detergent, and then used affinity chromatography and salt elution to isolate tubulin binding proteins. Dark-field microscopy of in vitro-assembled microtubules showed that the eluted proteins from both fractions induce microtubule bundling and, in the presence of purified tubullin, promote microtubule elongation. Gel electrophoresis of the eluted proteins revealed two distinct sets of polypeptides. Those in the membrane eluate included unique bands with apparent molecular masses of 98, 90, and 75 kD in addition to bands present in both eluates. The cytosolic eluate, in contrast, typically included relatively smaller proteins. The eluted proteins also bound to taxol-stabilized microtubules. Initial immunological characterization using monoclonal antibodies raised against the 90-kD polypeptide showed that it is colocalized in situ with cortical microtubules in tobacco protoplast ghosts
ISSN:1040-4651
1532-298X
DOI:10.1105/tpc.8.11.2127