No abstract

In order to assess the possible role of microfilaments (Mfs) in events preceding plant cell division, actin was localized in root cells of Allium cepa and Tradescantia vivginiana by immunofluorescence microscopy. The distribution of Mfs was compared to that of microtubules (Mts) by means of dual localizations employing both antiactin and antitubulin. Cycling interphase cells contain Mfs that extend into all regions of the cytoplasm in random fashion. Prior to the rearrangement of the cortical Mt array into the initial broad preprophase band (PPB), the number of Mfs in the cytoplasm decreases, while a new population appears in the cortex. The cortical Mfs, which usually occupy the entire cell surface, are aligned parallel to the cortical Mts. When the initial PPB appears, these Mfs still cover the cortex or are arranged as a broad band encompassing the PPB. As the PPB narrows, the Mfs are also confined to an increasingly restricted zone usually wider than the PPB. When the PPB reaches its narrowest, densest configuration, aligned Mfs are excluded from the band proper, while others appear in flanking regions of the cortex. From prometaphase through anaphase, cortical Mfs are largely restricted to the ends of the cell overlying the spindle poles; they also tend to become more randomly oriented. Little or no actin is present in the spindle. During telophase, the two zones of aligned cortical Mfs over the ends of the cell gradually disappear and are replaced by new interphase networks. These changes provide additional data on the possible control of PPB organization by actin, and in addition indicate that the cortex may be the origin of the actin that aggregates at the spindle poles during cytochalasin treatment.1992 Wiley-Liss, Inc.

Lucifer yellow has been microinjected into stomatal cells of Allium cepa L. epidermal slices and Commelina communis L. epidermal peels and the symplastic spread of dye to neighboring cells monitored by fluorescence microscopy. Dye does not move out of injected mature guard cells, nor does it spread into the guard cells when adjacent epidermal or subsidiary cells are injected. Dye does spread from injected subsidiary cells to other subsidiary cells. These results are consistent with the reported absence of plasmodesmata in the walls of mature guard cells. Microinjection was also used to ascertain when dye coupling ceases during stomatal differentiation in Allium. Dye rapidly moves into and out of guard mother cells and young guard cells. Hovewer, dye movement ceases midway through development as the guard cells begin to swell but well before a pore first opens. Since plasmodesmata are still present at this stage, the loss of symplastic transport may result from changes in these structures well in advance of their actual disappearance from the guard cell wall.

gamma-Tubulin is a protein associated with microtubule (Mt)-organizing centers in a variety of eukaryotic cells. Unfortunately, little is known about such centers in plants. Genomic and partial cDNA clones encoding two gamma-tubulins of Arabidopsis were isolated and sequenced. Comparisons of genomic and cDNA sequences showed that both genes, TubG1 and TubG2, contain nine introns at conserved locations. The sequences of the two genes both predict proteins containing 474 amino acids, with molecular masses of 53,250 and 53,280 D, respectively. The predicted gamma 1- and gamma 2-tubulins exhibit 98% amino acid identity with each other and approximately 70% amino acid identity with the gamma-tubulins of animals and fungi. RNA gel blot results demonstrated that both genes are transcribed in suspension culture cells, seedlings, and roots and flowers of mature plants. Immunoblots of Arabidopsis proteins using an antibody specific to a conserved peptide of gamma-tubulin showed a major cross-reacting polypeptide with an M(r) of 58,000. The same antibody stained all Mt arrays in tissue and suspension culture cells of this species. Binding was inhibited by the homologous oligopeptide in the gamma-tubulins predicted by the two Arabidopsis gene sequences. Antibody staining avoided the plus ends of Mts at the kinetochores and cell plate, but unlike the case in animal cells, seemed to be localized over broad stretches of the kinetochore fibers and phragmoplast toward the minus ends. We concluded that at least two gamma-tubulin protein homologs are present in Arabidopsis and that at least one of them is localized along Mt arrays. Its distribution is correlated with and may help explain unique characteristics of Mt organization in plants.

y-Tubulin is a protein associated with microtubule (Mt)-organizing centers in a variety of eukaryotic cells. Unfortunately, little is known about such centers in plants. Genomic and partia1 cDNA clones encoding two y-tubulins of Arabidopsis were isolated and sequenced. Comparisons of genomic and cDNA sequences showed that both genes, TubGl and TubGP, contain nine introns at conserved locations. The sequences of the two genes both predict proteins containing 474 amino acids, with molecular masses of 53,250 and 53,280 D, respectively. The predicted yl-and y2-tubulins exhibit 98% amino acid identity with each other and -70% amino acid identity with the y-tubulins of animals and fungi. RNA gel blot results demonstrated that both genes are transcribed in suspension culture cells, seedlings, and roots and flowers of mature plants. lmmunoblots of Arabidopsis proteins using an antibody specific to a conserved peptide of y-tubulin showed a major cross-reacting polypeptide with an M, of 58,000. The same antibody stained all Mt arrays in tissue and suspension culture cells of this species. Binding was inhibited by the homologous oligopeptide in the y-tubulins predicted by the two Arabidopsis gene sequences. Antibody staining avoided the plus ends of Mts at the kinetochores and cell plate, but unlike the case in animal cells, seemed to be localized over broad stretches of the kinetochore fibers and phragmoplast toward the minus ends. We concluded that at least two y-tubulin protein homologs are present in Arabidopsis and that at least one of them is localized along Mt arrays. Its distribution is correlated with and may help explain unique characteristics of Mt organization in plants.