scholarly journals Essential Roles for GPI-anchored Proteins in African Trypanosomes Revealed Using Mutants Deficient in GPI8

2003 ◽  
Vol 14 (3) ◽  
pp. 1182-1194 ◽  
Author(s):  
Simon Lillico ◽  
Mark C. Field ◽  
Pat Blundell ◽  
Graham H. Coombs ◽  
Jeremy C. Mottram
Keyword(s):  
Trypanosoma Brucei ◽  
Cell Cycle Progression ◽  
Tsetse Fly ◽  
Bloodstream Form ◽  
Surface Glycoprotein ◽  
Surface Coat ◽  
Immune Challenge ◽  
Gene Encoding ◽  
Cycle Progression ◽  
African Trypanosomes

The survival of Trypanosoma brucei, the causative agent of Sleeping Sickness and Nagana, is facilitated by the expression of a dense surface coat of glycosylphosphatidylinositol (GPI)-anchored proteins in both its mammalian and tsetse fly hosts. We have characterized T. brucei GPI8, the gene encoding the catalytic subunit of the GPI:protein transamidase complex that adds preformed GPI anchors onto nascent polypeptides. Deletion ofGPI8 (to give Δgpi8) resulted in the absence of GPI-anchored proteins from the cell surface of procyclic form trypanosomes and accumulation of a pool of non–protein-linked GPI molecules, some of which are surface located. Procyclic Δgpi8, while viable in culture, were unable to establish infections in the tsetse midgut, confirming that GPI-anchored proteins are essential for insect-parasite interactions. Applying specific inducible GPI8 RNAi with bloodstream form parasites resulted in accumulation of unanchored variant surface glycoprotein and cell death with a defined multinuclear, multikinetoplast, and multiflagellar phenotype indicative of a block in cytokinesis. These data show that GPI-anchored proteins are essential for the viability of bloodstream form trypanosomes even in the absence of immune challenge and imply that GPI8 is important for proper cell cycle progression.

Maintaining the protective variant surface glycoprotein coat of African trypanosomes

2005 ◽  
Vol 33 (5) ◽  
pp. 981-982 ◽  
Author(s):  
G. Rudenko
Keyword(s):  
Cell Cycle ◽  
Trypanosoma Brucei ◽  
Chronic Infection ◽  
Cell Cycle Progression ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
Cycle Progression ◽  
African Trypanosomes ◽  
Immune Attack ◽  
Protective Variant

The African trypanosome Trypanosoma brucei has a precarious existence as an extracellular parasite of the mammalian bloodstream, where it is faced with continuous immune attack. Key to survival is a dense VSG (variant surface glycoprotein) coat, which is repeatedly switched during the course of a chronic infection. New data demonstrate a link between VSG synthesis and cell cycle progression, indicating that VSG is monitored during the trypanosome cell cycle.

scholarly journals Escaping the immune system by DNA repair and recombination in African trypanosomes

Open Biology ◽  
2019 ◽  
Vol 9 (11) ◽  
pp. 190182 ◽  
Author(s):  
Núria Sima ◽  
Emilia Jane McLaughlin ◽  
Sebastian Hutchinson ◽  
Lucy Glover
Keyword(s):  
Immune Response ◽  
Dna Repair ◽  
Trypanosoma Brucei ◽  
Antigenic Variation ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
Surface Coat ◽  
African Trypanosomes ◽  
Active Expression ◽  
Expression Site

African trypanosomes escape the mammalian immune response by antigenic variation—the periodic exchange of one surface coat protein, in Trypanosoma brucei the variant surface glycoprotein (VSG), for an immunologically distinct one. VSG transcription is monoallelic, with only one VSG being expressed at a time from a specialized locus, known as an expression site. VSG switching is a predominantly recombination-driven process that allows VSG sequences to be recombined into the active expression site either replacing the currently active VSG or generating a ‘new’ VSG by segmental gene conversion. In this review, we describe what is known about the factors that influence this process, focusing specifically on DNA repair and recombination.

scholarly journals Expression site attenuation mechanistically links antigenic variation and development in Trypanosoma brucei

eLife ◽  
2014 ◽  
Vol 3 ◽  
Author(s):  
Christopher Batram ◽  
Nicola G Jones ◽  
Christian J Janzen ◽  
Sebastian M Markert ◽  
Markus Engstler
Keyword(s):  
Trypanosoma Brucei ◽  
Histone Methylation ◽  
Antigenic Variation ◽  
Tsetse Fly ◽  
Developmental Competence ◽  
Surface Glycoprotein ◽  
G1 Phase ◽  
African Trypanosomes ◽  
Developmental Progression ◽  
Expression Site

We have discovered a new mechanism of monoallelic gene expression that links antigenic variation, cell cycle, and development in the model parasite Trypanosoma brucei. African trypanosomes possess hundreds of variant surface glycoprotein (VSG) genes, but only one is expressed from a telomeric expression site (ES) at any given time. We found that the expression of a second VSG alone is sufficient to silence the active VSG gene and directionally attenuate the ES by disruptor of telomeric silencing-1B (DOT1B)-mediated histone methylation. Three conserved expression-site-associated genes (ESAGs) appear to serve as signal for ES attenuation. Their depletion causes G1-phase dormancy and reversible initiation of the slender-to-stumpy differentiation pathway. ES-attenuated slender bloodstream trypanosomes gain full developmental competence for transformation to the tsetse fly stage. This surprising connection between antigenic variation and developmental progression provides an unexpected point of attack against the deadly sleeping sickness.

scholarly journals Active VSG Expression Sites in Trypanosoma brucei Are Depleted of Nucleosomes

2009 ◽  
Vol 9 (1) ◽  
pp. 136-147 ◽  
Author(s):  
Tara M. Stanne ◽  
Gloria Rudenko
Keyword(s):  
Rna Polymerase ◽  
Trypanosoma Brucei ◽  
Histone H3 ◽  
Bloodstream Form ◽  
Surface Glycoprotein ◽  
Micrococcal Nuclease ◽  
Pcr Analysis ◽  
Marker Genes ◽  
African Trypanosomes ◽  
Pol I

ABSTRACT African trypanosomes regulate transcription differently from other eukaryotes. Most of the trypanosome genome is constitutively transcribed by RNA polymerase II (Pol II) as large polycistronic transcription units while the genes encoding the major surface proteins are transcribed by RNA polymerase I (Pol I). In bloodstream form Trypanosoma brucei, the gene encoding the variant surface glycoprotein (VSG) coat is expressed in a monoallelic fashion from one of about 15 VSG bloodstream form expression sites (BESs). Little is known about the chromatin structure of the trypanosome genome, and the chromatin state of active versus silent VSG BESs remains controversial. Here, we determined histone H3 occupancy within the genome of T. brucei, focusing on active versus silent VSG BESs in the bloodstream form. We found that histone H3 was most enriched in the nontranscribed 50-bp and 177-bp repeats and relatively depleted in Pol I, II, and III transcription units, with particular depletion over promoter regions. Using two isogenic T. brucei lines containing marker genes in different VSG BESs, we determined that histone H3 is 11- to 40-fold depleted from active VSG BESs compared with silent VSG BESs. Quantitative PCR analysis of fractionated micrococcal nuclease-digested chromatin revealed that the active VSG BES is depleted of nucleosomes. Therefore, in contrast to earlier views, nucleosome positioning appears to be involved in the monoalleleic control of VSG BESs in T. brucei. This may provide a level of epigenetic regulation enabling bloodstream form trypanosomes to efficiently pass on the transcriptional state of active and silent BESs to daughter cells.

scholarly journals Procyclin gene expression and loss of the variant surface glycoprotein during differentiation of Trypanosoma brucei.

1989 ◽  
Vol 108 (2) ◽  
pp. 737-746 ◽  
Author(s):  
I Roditi ◽  
H Schwarz ◽  
T W Pearson ◽  
R P Beecroft ◽  
M K Liu ◽  
...  
Keyword(s):  
Trypanosoma Brucei ◽  
Tsetse Fly ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
Mammalian Host ◽  
Cylindrical Shape ◽  
Surface Coat ◽  
Insect Vector ◽  
Molecular Arrangement

In the mammalian host, the unicellular flagellate Trypanosoma brucei is covered by a dense surface coat that consists of a single species of macromolecule, the membrane form of the variant surface glycoprotein (mfVSG). After uptake by the insect vector, the tsetse fly, bloodstream-form trypanosomes differentiate to procyclic forms in the fly midgut. Differentiation is characterized by the loss of the mfVSG coat and the acquisition of a new surface glycoprotein, procyclin. In this study, the change in surface glycoprotein composition during differentiation was investigated in vitro. After triggering differentiation, a rapid increase in procyclin-specific mRNA was observed. In contrast, there was a lag of several hours before procyclin could be detected. Procyclin was incorporated and uniformly distributed in the surface coat. The VSG coat was subsequently shed. For a single cell, it took 12-16 h to express a maximum level of procyclin at the surface while the loss of the VSG coat required approximately 4 h. The data are discussed in terms of the possible molecular arrangement of mfVSG and procyclin at the cell surface. Molecular modeling data suggest that a (Asp-Pro)2 (Glu-Pro)22-29 repeat in procyclin assumes a cylindrical shape 14-18 nm in length and 0.9 nm in diameter. This extended shape would enable procyclin to interdigitate between the mfVSG molecules during differentiation, exposing epitopes beyond the 12-15-nm-thick VSG coat.

scholarly journals Single-cell transcriptomic analysis of bloodstream Trypanosoma brucei reconstructs cell cycle progression and developmental quorum sensing

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Emma M. Briggs ◽  
Federico Rojas ◽  
Richard McCulloch ◽  
Keith R. Matthews ◽  
Thomas D. Otto
Keyword(s):  
Cell Cycle ◽  
Single Cell ◽  
Trypanosoma Brucei ◽  
Cell Cycle Progression ◽  
Expression Patterns ◽  
Tsetse Fly ◽  
Cycle Progression ◽  
Progressive Development ◽  
Asynchronous Development

AbstractDevelopmental steps in the trypanosome life-cycle involve transition between replicative and non-replicative forms specialised for survival in, and transmission between, mammalian and tsetse fly hosts. Here, using oligopeptide-induced differentiation in vitro, we model the progressive development of replicative ‘slender’ to transmissible ‘stumpy’ bloodstream form Trypanosoma brucei and capture the transcriptomes of 8,599 parasites using single cell transcriptomics (scRNA-seq). Using this framework, we detail the relative order of biological events during asynchronous development, profile dynamic gene expression patterns and identify putative regulators. We additionally map the cell cycle of proliferating parasites and position stumpy cell-cycle exit at early G1 before progression to a distinct G0 state. A null mutant for one transiently elevated developmental regulator, ZC3H20 is further analysed by scRNA-seq, identifying its point of failure in the developmental atlas. This approach provides a paradigm for the dissection of differentiation events in parasites, relevant to diverse transitions in pathogen biology.

scholarly journals TbSAP is a novel chromatin protein repressing metacyclic variant surface glycoprotein expression sites in bloodstream form Trypanosoma brucei

2021 ◽  
Vol 49 (6) ◽  
pp. 3242-3262
Author(s):  
Carys Davies ◽  
Cher-Pheng Ooi ◽  
Georgios Sioutas ◽  
Belinda S Hall ◽  
Haneesh Sidhu ◽  
...  
Keyword(s):  
Trypanosoma Brucei ◽  
Nuclear Periphery ◽  
Tsetse Fly ◽  
Bloodstream Form ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
Mammalian Host ◽  
Unicellular Eukaryote ◽  
Chromatin Protein ◽  
Protective Variant

Abstract The African trypanosome Trypanosoma brucei is a unicellular eukaryote, which relies on a protective variant surface glycoprotein (VSG) coat for survival in the mammalian host. A single trypanosome has >2000 VSG genes and pseudogenes of which only one is expressed from one of ∼15 telomeric bloodstream form expression sites (BESs). Infectious metacyclic trypanosomes present within the tsetse fly vector also express VSG from a separate set of telomeric metacyclic ESs (MESs). All MESs are silenced in bloodstream form T. brucei. As very little is known about how this is mediated, we performed a whole genome RNAi library screen to identify MES repressors. This allowed us to identify a novel SAP domain containing DNA binding protein which we called TbSAP. TbSAP is enriched at the nuclear periphery and binds both MESs and BESs. Knockdown of TbSAP in bloodstream form trypanosomes did not result in cells becoming more ‘metacyclic-like'. Instead, there was extensive global upregulation of transcripts including MES VSGs, VSGs within the silent VSG arrays as well as genes immediately downstream of BES promoters. TbSAP therefore appears to be a novel chromatin protein playing an important role in silencing the extensive VSG repertoire of bloodstream form T. brucei.

scholarly journals Tsetse Fly Saliva Accelerates the Onset of Trypanosoma brucei Infection in a Mouse Model Associated with a Reduced Host Inflammatory Response

2006 ◽  
Vol 74 (11) ◽  
pp. 6324-6330 ◽  
Author(s):  
Guy Caljon ◽  
Jan Van Den Abbeele ◽  
Benoît Stijlemans ◽  
Marc Coosemans ◽  
Patrick De Baetselier ◽  
...  
Keyword(s):  
Trypanosoma Brucei ◽  
Inflammatory Responses ◽  
Blood Feeding ◽  
Immunoglobulin M ◽  
Tsetse Fly ◽  
Surface Glycoprotein ◽  
Tsetse Flies ◽  
Feeding Site ◽  
African Trypanosomes ◽  
Cytokine Tumor Necrosis Factor

ABSTRACT Tsetse flies (Glossina sp.) are the vectors that transmit African trypanosomes, protozoan parasites that cause human sleeping sickness and veterinary infections in the African continent. These blood-feeding dipteran insects deposit saliva at the feeding site that enables the blood-feeding process. Here we demonstrate that tsetse fly saliva also accelerates the onset of a Trypanosoma brucei infection. This effect was associated with a reduced inflammatory reaction at the site of infection initiation (reflected by a decrease of interleukin-6 [IL-6] and IL-12 mRNA) as well as lower serum concentrations of the trypanocidal cytokine tumor necrosis factor. Variant-specific surface glycoprotein-specific antibody isotypes immunoglobulin M (IgM) and IgG2a, implicated in trypanosome clearance, were not suppressed. We propose that tsetse fly saliva accelerates the onset of trypanosome infection by inhibiting local and systemic inflammatory responses involved in parasite control.

scholarly journals Evidence for an interplay between cell cycle progression and the initiation of differentiation between life cycle forms of African trypanosomes.

1994 ◽  
Vol 125 (5) ◽  
pp. 1147-1156 ◽  
Author(s):  
K R Matthews ◽  
K Gull
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Blood Stream ◽  
Tsetse Fly ◽  
Mammalian Host ◽  
Cycle Progression ◽  
African Trypanosomes ◽  
Host Blood ◽  
Proliferative Cell ◽  
Differentiation Model

Successful transmission of the African trypanosome between the mammalian host blood-stream and the tsetse fly vector involves dramatic alterations in the parasite's morphology and biochemistry. This differentiation through to the tsetse midgut procyclic form is accompanied by re-entry into a proliferative cell cycle. Using a synchronous differentiation model and a variety of markers diagnostic for progress through both differentiation and the cell cycle, we have investigated the interplay between these two processes. Our results implicate a relationship between the trypanosome cell cycle position and the perception of the differentiation signal and demonstrate that irreversible commitment to the differentiation occurs rapidly after induction. Furthermore, we show that re-entry into the cell cycle in the differentiating population is synchronous, and that once initiated, progress through the differentiation pathway can be uncoupled from progress through the cell cycle.

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