A gamma-tubulin antibody against a plant peptide sequence localises to cell division-specific microtubule arrays and organelles in plants.
Gamma tubulin (gamma-tubulin) is involved in microtubule initiation in the eukaryotes. In animal cells it is localised to centrosomes and to other, non-centrosomal sites of microtubule initiation. In addition, cytoplasmic complexes containing gamma-tubulin (gamma-TuRCs; gamma-somes) have been described, which are multiprotein complexes involved in microtubule initiation. Most localisations of gamma-tubulin in plants have previously been achieved using an antibody directed towards a conserved peptide sequence found in animal cells, showing co-localisation with all microtubule arrays throughout the cell cycle. Because different antibodies may give various patterns of subcellular localisation, in the present study we raised a polyclonal antibody ('Hayley') to the plant peptide sequence EDFATQGGDRKDVFFY (bold letters indicate plant-specific amino acids) to further investigate the subcellular distribution in plants. Immunoblotting using wheat root tip protein extracts revealed a 58 kDagamma-tubulin-like peptide as has been described before. Immunofluorescence microscopy of wheat root-tip cells, however, revealed localisation of gamma-tubulin to a subset of mitotic microtubule arrays and the cytokinetic phragmoplast, but not to interphase cortical arrays or the preprophase band of microtubules. This lack of labelling may be caused by a restriction of antibody access during interphase, but more likely by a cell division-specific conformational change in the gamma-tubulin molecule. Our antibody also gave an organelle-like labelling, not described before, which may represent storage forms or precursors of gamma-tubulin, perhaps related to plastid-based microtubule initiation in hepatics and hornworts.
Dibbayawan
T P
TP
School of Biological Sciences, Macleay Building, A12, University of Sydney, NSW 2006, Sydney, Australia.
Harper
J D
JD
Marc
J
J
D000595:Amino Acid Sequence; D000818:Animals; D000906:Antibodies / Q000276:immunology*; D002455:Cell Division / Q000502:physiology; D018385:Centrosome / Q000737:chemistry; D019476:Insect Proteins; D018613:Microscopy, Confocal; D008856:Microscopy, Fluorescence; D008869:Microtubule-Associated Proteins / Q000032:analysis / Q000737:chemistry / Q000276:immunology; D022101:Microtubule-Organizing Center / Q000737:chemistry / Q000648:ultrastructure; D008870:Microtubules / Q000737:chemistry* / Q000648:ultrastructure; D008969:Molecular Sequence Data; D015388:Organelles / Q000737:chemistry* / Q000648:ultrastructure; D010455:Peptides / Q000737:chemistry; D018517:Plant Roots / Q000737:chemistry* / Q000648:ultrastructure; D011817:Rabbits; D014908:Triticum / Q000737:chemistry*; D014404:Tubulin / Q000032:analysis* / Q000737:chemistry / Q000276:immunology*
D000906:Antibodies; D019476:Insect Proteins; D008869:Microtubule-Associated Proteins; D010455:Peptides; D014404:Tubulin; C054628:microtubule-organizing center-associated polypeptide, insect