MICROTUBULE ORGANIZATION 1 regulates structure and function of microtubule arrays during mitosis and cytokinesis in the Arabidopsis root.

MICROTUBULE ORGANIZATION 1 (MOR1) is a plant member of the highly conserved MAP215/Dis1 family of microtubule-associated proteins. Prior studies with the temperature-sensitive mor1 mutants of Arabidopsis (Arabidopsis thaliana), which harbor single amino acid substitutions in an N-terminal HEAT repeat, proved that MOR1 regulates cortical microtubule organization and function. Here we demonstrate by use of live cell imaging and immunolabeling that the mor1-1 mutation generates specific defects in the microtubule arrays of dividing vegetative cells. Unlike the universal cortical microtubule disorganization in elongating mor1-1 cells, disruption of mitotic and cytokinetic microtubule arrays was not detected in all dividing cells. Nevertheless, quantitative analysis identified distinct defects in preprophase bands (PPBs), spindles, and phragmoplasts. In nearly one-half of dividing cells at the restrictive temperature of 30 degrees C, PPBs were not detected prior to spindle formation, and those that did form were often disrupted. mor1-1 spindles and phragmoplasts were short and abnormally organized and persisted for longer times than in wild-type cells. The reduced length of these arrays predicts that the component microtubule lengths are also reduced, suggesting that microtubule length is a critical determinant of spindle and phragmoplast structure, orientation, and function. Microtubule organizational defects led to aberrant chromosomal arrangements, misaligned or incomplete cell plates, and multinucleate cells. Antiserum raised against an N-terminal MOR1 sequence labeled the full length of microtubules in interphase arrays, PPBs, spindles, and phragmoplasts. Continued immunolabeling of the disorganized and short microtubules of mor1-1 at the restrictive temperature demonstrated that the mutant mor1-1(L174F) protein loses function without dissociating from microtubules, providing important insight into the mechanism by which MOR1 may regulate microtubule length.

Phylogenetic analysis of the erythrocytic Anaplasma species based on 16S rDNA and GroEL (HSP60) sequences of A. marginale, A. centrale, and A. ovis and the specific detection of A. centrale vaccine strain.

Phenotypic criteria for the identification of erythrocytic ruminant Anaplasma species has relied on subjective identification methods such as host pathogenicity (virulence for cattle or sheep) and/or the location of Anaplasma inclusion bodies within the host's red cells. Sequence comparisons of new and available GenBank Accessions were investigated to elucidate the relationships among these closely related Anaplasma species. Twenty-one 16S rDNA and GroEL (HSP60) sequences from 13 Anaplasma marginale (South Africa, Namibia, Zimbabwe, Israel, USA, Australia and Uruguay), three A. centrale (South Africa and Japan), two A. ovis (USA and South Africa), and two unknown Anaplasma species isolated from wild ruminants (South Africa), were compared. 16S rDNA maximum-likelihood and distance trees separated all A. marginale (and the two wild ruminant isolates) from the two South African A. centrale (including original vaccine strain, Theiler, 1911). The Japanese A. centrale (Aomori) demonstrated the lowest sequence identity to the remaining erythrocytic Anaplasma species. A. ovis inter-species relationships could not be resolved through the 16S rDNA analyses, whereas strong bootstrap branch support is demonstrated in the GroEL distance tree using A. ovis OVI strain. All erythrocytic Anaplasma species and isolates were confirmed to belong to the same cluster showing strong branch support to Anaplasma (Ehrlichia) phagocytophilum with Ehrlichia (Cowdria) ruminantium and Rickettsia rickettsii serving as appropriate out-groups. Based on groEL sequences, a specific PCR method was developed which amplified A. centrale vaccine (Theiler, 1911) specifically. This study confirms the suitability of 16S rDNA sequences to define genera and demonstrates the usefulness of GroEL sequences for defining species of erythrocytic Anaplasma.