This study uses electron tomography linked to a variety of other EM methods to provide an integrated view of the flagellar pocket and basal body area of the African trypanosome procyclic trypomastigote. We reveal the pocket as an asymmetric membranous `balloon' with two boundary structures. One of these – the collar – defines the flagellum exit point. The other defines the entry point of the flagellum into the pocket and consists of both an internal transitional fibre array and an external membrane collarette. A novel set of nine radial fibres is described in the basal body proximal zone. The pocket asymmetry is invariably correlated with the position of the probasal body and Golgi. The neck region, just distal to the flagellum exit site, is a specialised area of membrane associated with the start of the flagellum attachment zone and signifies the point where a special set of four microtubules, nucleated close to the basal bodies, joins the subpellicular array. The neck region is also associated with the single Golgi apparatus of the cell. The flagellar exit point interrupts the subpellicular microtubule array with discrete endings of microtubules at the posterior side. Overall, our studies reveal a highly organised, yet dynamic, area of cytoplasm and will be informative in understanding its function.

The FtsZ protein is a polymer-forming GTPase which drives bacterial cell division and is structurally and functionally related to eukaryotic tubulins. We have searched for FtsZ-related sequences in all freely accessible databases, then used strict criteria based on the tertiary structure of FtsZ and its well-characterized in vitro and in vivo properties to determine which sequences represent genuine homologues of FtsZ. We have identified 225 full-length FtsZ homologues, which we have used to document, phylum by phylum, the primary sequence characteristics of FtsZ homologues from the Bacteria, Archaea, and Eukaryota. We provide evidence for at least five independent ftsZ gene-duplication events in the bacterial kingdom and suggest the existence of three ancestoral euryarchaeal FtsZ paralogues. In addition, we identify "FtsZ-like" sequences from Bacteria and Archaea that, while showing significant sequence similarity to FtsZs, are unlikely to bind and hydrolyze GTP.

BackgroundPhotosynthetic euglenids are major contributors to fresh water ecosystems. Euglena gracilis in particular has noted metabolic flexibility, reflected by an ability to thrive in a range of harsh environments. E. gracilis has been a popular model organism and of considerable biotechnological interest, but the absence of a gene catalogue has hampered both basic research and translational efforts.ResultsWe report a detailed transcriptome and partial genome for E. gracilis Z1. The nuclear genome is estimated to be around 500 Mb in size, and the transcriptome encodes over 36,000 proteins and the genome possesses less than 1% coding sequence. Annotation of coding sequences indicates a highly sophisticated endomembrane system, RNA processing mechanisms and nuclear genome contributions from several photosynthetic lineages. Multiple gene families, including likely signal transduction components, have been massively expanded. Alterations in protein abundance are controlled post-transcriptionally between light and dark conditions, surprisingly similar to trypanosomatids.ConclusionsOur data provide evidence that a range of photosynthetic eukaryotes contributed to the Euglena nuclear genome, evidence in support of the ‘shopping bag’ hypothesis for plastid acquisition. We also suggest that euglenids possess unique regulatory mechanisms for achieving extreme adaptability, through mechanisms of paralog expansion and gene acquisition.Electronic supplementary materialThe online version of this article (10.1186/s12915-019-0626-8) contains supplementary material, which is available to authorized users.

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Trypanosoma brucei is a protozoan parasite that causes African sleeping sickness. T. brucei multiplies extracellularly in the bloodstream, relying on antigenic variation of a dense variant surface glycoprotein (VSG) coat to escape antibody-mediated lysis. We investigated the role of VSG in proliferation and pathogenicity by using inducible RNA interference to ablate VSG transcript down to 1-2% normal levels. Inhibiting VSG synthesis in vitro triggers a rapid and specific cell cycle checkpoint blocking cell division. Parasites arrest at a discrete precytokinesis stage with two fulllength flagella and opposing flagellar pockets, without undergoing additional rounds of S phase and mitosis. A subset (<10%) of the stalled cells have internal flagella, indicating that the progenitors of these cells were already committed to cytokinesis when VSG restriction was sensed. Although there was no obvious VSG depletion in vitro after 24-h induction of VSG RNA interference, there was rapid clearance of these cells in vivo. We propose that a stringent block in VSG synthesis produces stalled trypanosomes with a minimally compromised VSG coat, which can be targeted by the immune system. Our data indicate that VSG protein or transcript is monitored during cell cycle progression in bloodstreamform T. brucei and describes precise precytokinesis cell cycle arrest. This checkpoint before cell division provides a link between the protective VSG coat and cell cycle progression and could function as a novel parasite safety mechanism, preventing extensive dilution of the protective VSG coat in the absence of VSG synthesis. antigenic variation ͉ Trypanosoma bruceiA frican trypanosomes, including Trypanosoma brucei, are unicellular parasites transmitted by tsetse flies in subSaharan Africa, which are responsible for debilitating disease in humans and livestock. African sleeping sickness is resurgent, with an estimated 300,000-500,000 cases a year, and it is invariably fatal if untreated (1). T. brucei multiplies extracellularly in the bloodstream, exposing it to immune attack. Each bloodstream-form cell is coated with Ϸ10 7 variant surface glycoprotein (VSG) molecules of a given antigenic type, making up Ϸ10% of total cellular protein (2, 3). Eventually the host mounts an antibody response against a given VSG variant. However, as parasites can switch between hundreds of antigenically diverse VSG coats during the course of a chronic infection, trypanosomes expressing a new VSG variant can escape the host antibody response against the old one (4, 5). This highly sophisticated strategy of antigenic variation makes African trypanosomes a paradigm for antigenic variation in general.Although a great deal is known about mechanisms mediating VSG switching, we know relatively little about the role of VSG itself in other aspects of immune evasion and pathogenicity. To investigate this, we inhibited VSG synthesis by performing inducible RNA interference (RNAi) both in vitro and in vivo. Inhibition of VSG synthesis in vitro triggers a rapid and specifi...