MICROTUBULE POLARITY AND DYNAMICS IN THE CONTROL OF ORGANELLE POSITIONING, SEGREGATION, AND CYTOKINESIS IN THE TRYPANOSOME CELL-CYCLE

Trypanosoma brucei has a precisely ordered microtubule cytoskeleton wh ose morphogenesis is central to cell cycle events such as organelle po sitioning, segregation, mitosis, and cytokinesis. We have defined micr otubule polarity and show the + ends of the cortical microtubules to b e at the posterior end of the cell. Measurements of organelle position s through the cell cycle reveal a high degree of coordinate movement a nd a relationship with overall cell extension. Quantitative analysis o f the segregation of the replicated mitochondrial genome (the kinetopl ast) by the flagellar basal bodies identifies a new G2 cell cycle even t marker. The subsequent mitosis then positions one ''daughter'' nucle us into the gap between the segregated basal bodies/kinetoplasts. The anterior daughter nucleus maintains its position relative to the anter ior of the cell, suggesting an effective yet cryptic nuclear positioni ng mechanism. Inhibition of microtubule dynamics by rhizoxin results i n a phenomenon whereby cells, which have segregated their kinetoplasts yet are compromised in mitosis, cleave into a nucleated portion and a flagellated, anucleate, cytoplast. We term these cytoplasts ''zoids'' and show that they contain the posterior (new) flagellum and associat ed basal-body/ kinetoplast complex. Examination of zoids suggests a ro le for the flagellum attachment zone (FAZ) in defining the position fo r the axis of cleavage in trypanosomes. Progression through cytokinesi s, (zoid formation) while mitosis is compromised, suggests that the de pendency relationships leading to the classical cell cycle check point s may be altered in trypanosomes, to take account of the need to segre gate two unit genomes (nuclear and mitochondrial) in this cell.