TY - JOUR A1 - Zimmermann, Henriette A1 - Subota, Ines A1 - Batram, Christopher A1 - Kramer, Susanne A1 - Janzen, Christian J. A1 - Jones, Nicola G. A1 - Engstler, Markus T1 - A quorum sensing-independent path to stumpy development in Trypanosoma brucei JF - PLoS Pathogens N2 - For persistent infections of the mammalian host, African trypanosomes limit their population size by quorum sensing of the parasite-excreted stumpy induction factor (SIF), which induces development to the tsetse-infective stumpy stage. We found that besides this cell density-dependent mechanism, there exists a second path to the stumpy stage that is linked to antigenic variation, the main instrument of parasite virulence. The expression of a second variant surface glycoprotein (VSG) leads to transcriptional attenuation of the VSG expression site (ES) and immediate development to tsetse fly infective stumpy parasites. This path is independent of SIF and solely controlled by the transcriptional status of the ES. In pleomorphic trypanosomes varying degrees of ES-attenuation result in phenotypic plasticity. While full ES-attenuation causes irreversible stumpy development, milder attenuation may open a time window for rescuing an unsuccessful antigenic switch, a scenario that so far has not been considered as important for parasite survival. KW - Trypanosoma KW - hyperexpression techniques KW - parasitic cell cycles KW - cloning KW - cell cycle and cell division KW - cell differentiation KW - tetracyclines KW - parasitic diseases Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-158230 VL - 13 IS - 4 ER - TY - JOUR A1 - Schuster, Sarah A1 - Krüger, Timothy A1 - Subota, Ines A1 - Thusek, Sina A1 - Rotureau, Brice A1 - Beilhack, Andreas A1 - Engstler, Markus T1 - Developmental adaptations of trypanosome motility to the tsetse fly host environments unravel a multifaceted in vivo microswimmer system JF - eLife N2 - The highly motile and versatile protozoan pathogen Trypanosoma brucei undergoes a complex life cycle in the tsetse fly. Here we introduce the host insect as an expedient model environment for microswimmer research, as it allows examination of microbial motion within a diversified, secluded and yet microscopically tractable space. During their week-long journey through the different microenvironments of the fly´s interior organs, the incessantly swimming trypanosomes cross various barriers and confined surroundings, with concurrently occurring major changes of parasite cell architecture. Multicolour light sheet fluorescence microscopy provided information about tsetse tissue topology with unprecedented resolution and allowed the first 3D analysis of the infection process. High-speed fluorescence microscopy illuminated the versatile behaviour of trypanosome developmental stages, ranging from solitary motion and near-wall swimming to collective motility in synchronised swarms and in confinement. We correlate the microenvironments and trypanosome morphologies to high-speed motility data, which paves the way for cross-disciplinary microswimmer research in a naturally evolved environment. KW - none KW - tsetse fly KW - Trypanosoma KW - biophysics KW - microswimmer KW - sleeping sickness KW - structural biology Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-158662 VL - 6 ER - TY - JOUR A1 - Kramer, Susanne T1 - The ApaH-like phosphatase TbALPH1 is the major mRNA decapping enzyme of trypanosomes JF - PLoS Pathogens N2 - 5’-3’ decay is the major mRNA decay pathway in many eukaryotes, including trypanosomes. After deadenylation, mRNAs are decapped by the nudix hydrolase DCP2 of the decapping complex and finally degraded by the 5’-3’ exoribonuclease. Uniquely, trypanosomes lack homologues to all subunits of the decapping complex, while deadenylation and 5’-3’ degradation are conserved. Here, I show that the parasites use an ApaH-like phosphatase (ALPH1) as their major mRNA decapping enzyme. The protein was recently identified as a novel trypanosome stress granule protein and as involved in mRNA binding. A fraction of ALPH1 co-localises exclusively with the trypanosome 5’-3’ exoribonuclease XRNA to a special granule at the posterior pole of the cell, indicating a connection between the two enzymes. RNAi depletion of ALPH1 is lethal and causes a massive increase in total mRNAs that are deadenylated, but have not yet started 5’-3’ decay. These data suggest that ALPH1 acts downstream of deadenylation and upstream of mRNA degradation, consistent with a function in mRNA decapping. In vitro experiments show that recombinant, N-terminally truncated ALHP1 protein, but not a catalytically inactive mutant, sensitises the capped trypanosome spliced leader RNA to yeast Xrn1, but only if an RNA 5’ polyphosphatase is included. This indicates that the decapping mechanism of ALPH1 differs from the decapping mechanism of Dcp2 by leaving more than one phosphate group at the mRNA’s 5’ end. This is the first reported function of a eukaryotic ApaH-like phosphatase, a bacterial-derived class of enzymes present in all phylogenetic super-groups of the eukaryotic kingdom. The substrates of eukaryotic ApaH-like phosphatases are unknown. However, the substrate of the related bacterial enzyme ApaH, diadenosine tetraphosphate, is highly reminiscent of a eukaryotic mRNA cap. KW - eukaryota KW - Trypanosoma KW - RNA interference KW - messenger RNA Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-158482 VL - 13 IS - 6 ER - TY - JOUR A1 - Goos, Carina A1 - Dejung, Mario A1 - Janzen, Christian J. A1 - Butter, Falk A1 - Kramer, Susanne T1 - The nuclear proteome of Trypanosoma brucei JF - PLoS ONE N2 - Trypanosoma brucei is a protozoan flagellate that is transmitted by tsetse flies into the mammalian bloodstream. The parasite has a huge impact on human health both directly by causing African sleeping sickness and indirectly, by infecting domestic cattle. The biology of trypanosomes involves some highly unusual, nuclear-localised processes. These include polycistronic transcription without classical promoters initiated from regions defined by histone variants, trans-splicing of all transcripts to the exon of a spliced leader RNA, transcription of some very abundant proteins by RNA polymerase I and antigenic variation, a switch in expression of the cell surface protein variants that allows the parasite to resist the immune system of its mammalian host. Here, we provide the nuclear proteome of procyclic Trypanosoma brucei, the stage that resides within the tsetse fly midgut. We have performed quantitative label-free mass spectrometry to score 764 significantly nuclear enriched proteins in comparison to whole cell lysates. A comparison with proteomes of several experimentally characterised nuclear and non-nuclear structures and pathways confirmed the high quality of the dataset: the proteome contains about 80% of all nuclear proteins and less than 2% false positives. Using motif enrichment, we found the amino acid sequence KRxR present in a large number of nuclear proteins. KRxR is a sub-motif of a classical eukaryotic monopartite nuclear localisation signal and could be responsible for nuclear localization of proteins in Kinetoplastida species. As a proof of principle, we have confirmed the nuclear localisation of six proteins with previously unknown localisation by expressing eYFP fusion proteins. While proteome data of several T. brucei organelles have been published, our nuclear proteome closes an important gap in knowledge to study trypanosome biology, in particular nuclear-related processes. KW - Trypanosoma KW - gambiense KW - Trypanosoma brucei KW - proteomes KW - yellow fluorescent protein KW - mitochondria KW - protein structure KW - histones Y1 - 2017 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:bvb:20-opus-158572 VL - 12 IS - 7 ER -