Plant gamma-tubulin interacts with alphabeta-tubulin dimers and forms membrane-associated complexes.
gamma-Tubulin is assumed to participate in microtubule nucleation in acentrosomal plant cells, but the underlying molecular mechanisms are still unknown. Here, we show that gamma-tubulin is present in protein complexes of various sizes and different subcellular locations in Arabidopsis and fava bean. Immunoprecipitation experiments revealed an association of gamma-tubulin with alphabeta-tubulin dimers. gamma-Tubulin cosedimented with microtubules polymerized in vitro and localized along their whole length. Large gamma-tubulin complexes resistant to salt treatment were found to be associated with a high-speed microsomal fraction. Blue native electrophoresis of detergent-solubilized microsomes showed that the molecular mass of the complexes was >1 MD. Large gamma-tubulin complexes were active in microtubule nucleation, but nucleation activity was not observed for the smaller complexes. Punctate gamma-tubulin staining was associated with microtubule arrays, accumulated with short kinetochore microtubules interacting in polar regions with membranes, and localized in the vicinity of nuclei and in the area of cell plate formation. Our results indicate that the association of gamma-tubulin complexes with dynamic membranes might ensure the flexibility of noncentrosomal microtubule nucleation. Moreover, the presence of other molecular forms of gamma-tubulin suggests additional roles for this protein species in microtubule organization.
Dryková
Denisa
D
Institute of Microbiology, Academy of Sciences of the Czech Republic, Vídeská 1083, 142 20 Prague 4, Czech Republic.
Cenklová
Vēra
V
Sulimenko
Vadym
V
Volc
Jindrich
J
Dráber
Pavel
P
Binarová
Pavla
P
D000974:Antibodies, Antinuclear / Q000235:genetics / Q000378:metabolism; D017360:Arabidopsis / Q000378:metabolism*; D029681:Arabidopsis Proteins / Q000378:metabolism; D002462:Cell Membrane / Q000378:metabolism*; D003600:Cytosol / Q000378:metabolism; D019281:Dimerization; D004591:Electrophoresis, Polyacrylamide Gel; D005455:Fluorescent Antibody Technique; D008861:Microsomes / Q000378:metabolism; D008870:Microtubules / Q000378:metabolism; D008938:Mitosis / Q000502:physiology; D011233:Precipitin Tests; D011485:Protein Binding; D014404:Tubulin / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D031307:Vicia faba / Q000378:metabolism*
D000974:Antibodies, Antinuclear; D029681:Arabidopsis Proteins; D014404:Tubulin