Katanins and Spastin in Trypanosome Cytokinesis Figure 3. Depletion of KAT60a, KAT60b or KAT60c delays cytokinesis at an early stage in bloodstream trypanosomes. SEM images of RNAi cells (A: KAT60a; B: KAT60b; C: KAT60c) with 2 full length flagella. Cells were induced with tetracycline for 12.5 hours. Arrowheads indicate cleavage folds. Scale bars: 2 mm. doi:10.1371/journal.pone.0030367.g003 different stages, suggesting that the coordinated action of several microtubule severing enzymes is required to divide the trypano- some cytoskeleton. The KAT60 proteins appear to be important early on in cytokinesis, since the majority of 2N2K cells that accumulated following their depletion did not display cleavage furrows. Our data suggest that KAT60a may be important for conversion of the cleavage cleft to a furrow to separate the two daughter cell bodies. KAT60b on the other hand appears to be important for cleavage of the cytoskeleton at the anterior of the cell, since following its depletion, cells were observed with defined (although not separated) posterior daughter cell ends despite the lack of furrow ingression from the anterior. KAT60c is perhaps important at multiple stages of cytokinesis as no dominant morphological phenotype was observed amongst the 2N2K/2F cell population. Depletion of KAT80 and SPA affected later stages of cytokinesis, specifically inhibiting furrow ingression and abscission, respectively, as evidenced by the increased proportion of 2N2K cells with a partially ingressed cleavage furrow, or just joined at their posterior tips. One possible caveat in this interpretation of our data is that it is possible that the degree of protein knockdown induced by the RNAi for each of these severing proteins influences the stage of cytokinesis affected. For example, KAT80 and/or spastin could be required throughout cytokinesis, but in lower amounts during the early stages, and PLoS ONE | www.plosone.org 6 January 2012 | Volume 7 | Issue 1 | e30367