Higher plants represent a large group of eukaryotes where centrosomes are absent. The functions of γ-tubulin small complexes (γ-TuSCs) and γ-tubulin ring complexes (γ-TuRCs) in metazoans and fungi in microtubule nucleation are well established and the majority of components found in the complexes are present in plants. However, plant microtubules are also nucleated in a γ-tubulin-dependent but γ-TuRC-independent manner. There is growing evidence that γ-tubulin is a microtubule nucleator without being complexed in γ-TuRC. Fibrillar arrays of γ-tubulin were demonstrated in plant and animal cells and the ability of γ-tubulin to assemble into linear oligomers/polymers was confirmed in vitro for both native and recombinant γ-tubulin. The functions of γ-tubulin as a template for microtubule nucleation or in promoting spontaneous nucleation is outlined. Higher plants represent an excellent model for studies on the role of γ-tubulin in nucleation due to their acentrosomal nature and high abundancy and conservation of γ-tubulin including its intrinsic ability to assemble filaments. The defining scaffolding or sequestration functions of plant γ-tubulin in microtubule organization or in nuclear processes will help our understanding of its cellular roles in eukaryotes.

• Klíčová slova
• fibrillar arrays, gamma-tubulin, gamma-tubulin complexes, microtubules, nucleation, plants, sequestration, signaling,
• MeSH
• buňky metabolismus   MeSH
• centrozom metabolismus   MeSH
• lidé MeSH
• rostliny metabolismus   MeSH
• sekvence aminokyselin MeSH
• tubulin chemie   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• tubulin MeSH

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.

• MeSH
• antinukleární protilátky genetika   metabolismus   MeSH
• Arabidopsis metabolismus   MeSH
• buněčná membrána metabolismus   MeSH
• cytosol metabolismus   MeSH
• dimerizace MeSH
• elektroforéza v polyakrylamidovém gelu MeSH
• fluorescenční protilátková technika MeSH
• mikrotubuly metabolismus   MeSH
• mikrozomy metabolismus   MeSH
• mitóza fyziologie   MeSH
• precipitinové testy MeSH
• proteiny huseníčku metabolismus   MeSH
• tubulin chemie   imunologie   metabolismus   MeSH
• vazba proteinů MeSH
• Vicia faba metabolismus   MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• antinukleární protilátky MeSH
• proteiny huseníčku MeSH
• tubulin MeSH

gamma-Tubulin is necessary for nucleation and polar orientation of microtubules in vivo. The molecular mechanism of microtubule nucleation by gamma-tubulin and the regulation of this process are not fully understood. Here we show that there are two gamma-tubulin forms in the brain that are present in complexes of various sizes. Large complexes tend to dissociate in the presence of a high salt concentration. Both gamma-tubulins co-polymerized with tubulin dimers, and multiple gamma-tubulin bands were identified in microtubule protein preparations under conditions of non-denaturing electrophoresis. Immunoprecipitation experiments with monoclonal antibodies against gamma-tubulin and alpha-tubulin revealed interactions of both gamma-tubulin forms with tubulin dimers, irrespective of the size of complexes. We suggest that, besides small and large gamma-tubulin complexes, other molecular gamma-tubulin form(s) exist in brain extracts. Two-dimensional electrophoresis revealed multiple charge variants of gamma-tubulin in both brain extracts and microtubule protein preparations. Post-translational modification(s) of gamma-tubulins might therefore have an important role in the regulation of microtubule nucleation in neuronal cells.

• MeSH
• dimerizace MeSH
• elektroforéza v polyakrylamidovém gelu MeSH
• frakcionace buněk MeSH
• mozek - chemie * MeSH
• prasata MeSH
• protein - isoformy MeSH
• tkáňové extrakty chemie   metabolismus   MeSH
• tubulin chemie   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• protein - isoformy MeSH
• tkáňové extrakty MeSH
• tubulin MeSH