Microtubules, but not actin filaments, drive daughter cell budding and cell division in Toxoplasma gondii

We have used drugs to examine the role(s) of the actin and microtubule cyto skeletons in the intracellular growth and replication of the intracellular protozoan parasite, Toxoplasma gondii, By using a 5 minute infection period and adding the drugs shortly after entry we can treat parasites at the sta rt of intracellular development and 6-8 hours prior to the onset of daughte r cell budding, Using this approach we found, somewhat surprisingly, that r eagents that perturb the actin cytoskeleton in different ways (cytochalasin D, latrunculin A and jasplakinolide) had little effect on parasite replica tion although they had the expected effects on the host cells. These actin inhibitors did, however, disrupt the orderly turnover of the mother cell or ganelles lending to the formation of a large residual body at the posterior end of each pair of budding parasites. Treating established parasite cultu res with the actin inhibitors blocked ionophore-induced egression of tachyz oites from the host cells, demonstrating that intracellular parasites were susceptible to the effects of these inhibitors. In contrast, the anti-micro tubule drugs oryzalin and taxol, and to a much lesser extent nocodazole, wh ich affect microtubule dynamics in different ways, blocked parasite replica tion by disrupting the normal assembly of the apical conoid and the microtu bule inner membrane complex (IMC) in the budding daughter parasites. Centro some replication and assembly of intranuclear spindles, however, occurred n ormally. Thus, daughter cell budding per se is dependent primarily on the p arasite microtubule system and does not require a dynamic actin cytoskeleto n, although disruption of actin dynamics causes problems in the turnover of parasite organelles.