ABSTRACT
We describe here a new alpha-tubulin isoform from sunflower we named alpha(pi)-tubulin. Alpha(pi)-tubulin is the most divergent higher-plant alpha-tubulin described so far, having an unusual deletion in the H1/B2 loop and a glutamine-rich C-terminus. We constructed a three-dimensional model and discuss its implications. Using specific antibodies, we show that alpha(pi)-tubulin expression is restricted to the male gametophyte. Alpha(pi)-tubulin mRNA represents 90% of alpha-tubulin mRNA and a small percentage of total pollen mRNA. Among the plants tested, alpha(pi)-tubulin was only detected in sunflower and in Cosmos. Since both plants are Asteraceae, we propose that alpha(pi)-tubulin is specific to this family. Our results suggest that alpha(pi)-tubulin can inhibit tubulin assembly in pollen. This hypothesis is reinforced by the fact that alpha(pi)-tubulin is found in a complex with beta-tubulin in mature sunflower pollen.
Subject(s)
Helianthus/genetics , Pollen/genetics , Tubulin/genetics , Amino Acid Sequence , Asteraceae/genetics , Base Sequence , Blotting, Western , DNA, Complementary/chemistry , DNA, Complementary/genetics , Gene Expression , Immunohistochemistry , Microtubules/metabolism , Models, Molecular , Molecular Sequence Data , Pollen/growth & development , Protein Conformation , Protein Isoforms/chemistry , Protein Isoforms/genetics , Protein Isoforms/metabolism , Sequence Alignment , Sequence Analysis, DNA , Sequence Homology, Amino Acid , Nicotiana/genetics , Tubulin/chemistry , Tubulin/metabolismABSTRACT
The molecular basis of microtubule nucleation is still not known in higher plant cells. This process is better understood in yeast and animals cells. In the yeast spindle pole body and the centrosome in animal cells, gamma-tubulin small complexes and gamma-tubulin ring complexes, respectively, nucleate all microtubules. In addition to gamma-tubulin, Spc98p or its homologues plays an essential role. We report here the characterization of rice and Arabidopsis homologues of SPC98. Spc98p colocalizes with gamma-tubulin at the nuclear surface where microtubules are nucleated on isolated tobacco nuclei and in living cells. AtSpc98p-GFP also localizes at the cell cortex. Spc98p is not associated with gamma-tubulin along microtubules. These data suggest that multiple microtubule-nucleating sites are active in plant cells. Microtubule nucleation involving Spc98p-containing gamma-tubulin complexes could then be conserved among all eukaryotes, despite differences in structure and spatial distribution of microtubule organizing centers.
