A Trypanosoma brucei protein required for maintenance of the flagellum attachment zone and flagellar pocket ER domains

Trypanosomes and Leishmanias are important human parasites whose cellular architecture is centred on the single flagellum. In trypanosomes, this flagellum is attached to the cell along a complex flagellum attachment zone (FAZ), comprising flagellar and cytoplasmic components, the integrity of which is required for correct cell morphogenesis and division. The cytoplasmic FAZ cytoskeleton is conspicuously associated with a sheet of endoplasmic reticulum termed the 'FAZ ER'. In the present work, 3D electron tomography of bloodstream form trypanosomes was used to clarify the nature of the FAZ ER. We also identified TbVAP, a T. brucei protein whose knockdown by RNAi in procyclic form cells leads to a dramatic reduction in the FAZ ER, and in the ER associated with the flagellar pocket. TbVAP is an orthologue of VAMP-associated proteins (VAPs), integral ER membrane proteins whose mutation in humans has been linked to familial motor neuron disease. The localisation of tagged TbVAP and the phenotype of TbVAP RNAi in procyclic form trypanosomes are consistent with a function for TbVAP in the maintenance of sub-populations of the ER associated with the FAZ and the flagellar pocket. Nevertheless, depletion of TbVAP did not affect cell viability or cell cycle progression.