Functional analyses of the CIF1-CIF2 complex in Trypanosoma brucei identify the structural motifs required for complex formation and cytokinesis

Abstract

Cytokinesis in trypanosome occurs uni-directionally along the longitudinal axis from the cell anterior towards the cell posterior and requires a trypanosome-specific CIF1-CIF2 protein complex. However, little is known about the contribution of the structural motifs in CIF1 and CIF2 to complex assembly and cytokinesis. Here, we demonstrated that the two zinc-finger motifs but not the coiled-coil motif in CIF1 are required for interaction with the EF-hand motifs in CIF2. We further showed that localization of CIF1 depends on the coiled-coil motif and the first zinc-finger motif and that localization of CIF2 depends on the EF-hand motifs. Deletion of the coiled-coil motif and mutation of either zinc-finger motifs in CIF1 disrupted cytokinesis. Further, mutation of either zinc-finger motif in CIF1 mis-localized CIF2 to the cytosol and destabilized CIF2, whereas deletion of the coiled-coil motif in CIF1 spread CIF2 over to the new flagellum attachment zone and stabilized CIF2. Together, these results uncovered the requirement of the coiled-coil motif and zinc-finger motifs for CIF1 function in cytokinesis and for CIF2 localization and stability, providing structural insights into the functional interplay between the two cytokinesis regulators.