Abstract
Trypanosoma brucei, a flagellated protozoan parasite causing human sleeping sickness, relies on a subpellicular microtubule array for maintenance of cell morphology. The flagellum is attached to the cell body through a poorly understood flagellum attachment zone (FAZ), and regulates cell morphogenesis using an unknown mechanism. Here we identified a new FAZ component, CC2D, which contains coiled-coil motifs followed by a C-terminal C2 domain. T. brucei CC2D is present on the FAZ filament, FAZ-juxtaposed ER membrane and the basal bodies. Depletion of CC2D inhibits the assembly of a new FAZ filament, forming a FAZ stub with a relatively fixed size at the base of a detached, but otherwise normal, flagellum. Inhibition of new FAZ formation perturbs subpellicular microtubule organization and generates short daughter cells. The cell length shows a strong linear correlation with FAZ length, in both control cells and in cells with inhibited FAZ assembly. Together, our data support a direct function of FAZ assembly in determining new daughter cell length by regulating subpellicular microtubule synthesis.
Highlights
Trypanosoma brucei is a protozoan parasite containing a number of single-copy organelles encaged in a subpellicular microtubule array, all have to be duplicated and partitioned into the daughter cells during the cell cycle (Vaughan and Gull, 2008)
We report a new flagellum attachment zone (FAZ) marker CC2D that is present on the basal bodies and FAZ in procyclic T. brucei
T. brucei CC2D In a previous comparative proteomic study that compared the protein compositions between isolated flagella and flagellar complex (Zhou et al, 2010), one candidate protein (GeneID: Tb927.4.2080) was found associated with the flagellum or the FAZ when transiently overexpressed as a fusion to yellow fluorescent protein (YFP)
Summary
Trypanosoma brucei is a protozoan parasite containing a number of single-copy organelles encaged in a subpellicular microtubule array, all have to be duplicated and partitioned into the daughter cells during the cell cycle (Vaughan and Gull, 2008). The FAZ is characterized by an electron-dense filament structure subtending the flagellum and a set of four microtubules in close association with smooth ER membranes (Sherwin and Gull, 1989a; Vickerman 1962; Vickerman, 1969). In procyclic T. brucei, new FAZ filament assembles together with the elongation of the new flagellum during the cell cycle (Kohl et al, 1999), always posterior to the old structures. With the cleavage furrow ingressing posteriorly, the duplicated organelles are partitioned into two daughter cells, one inherits the old, more anteriorly located FAZ and flagellum and the other inherits the newly synthesized, more posteriorly located FAZ and flagellum
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