scholarly journals To the Surface and Back: Exo- and Endocytic Pathways in Trypanosoma brucei

2021 ◽  
Vol 9 ◽  
Author(s):  
Fabian Link ◽  
Alyssa R. Borges ◽  
Nicola G. Jones ◽  
Markus Engstler
Keyword(s):  
Cell Surface ◽  
Trypanosoma Brucei ◽  
Immune Evasion ◽  
Antigenic Variation ◽  
Critical Role ◽  
Surface Glycoprotein ◽  
Immune Recognition ◽  
Flagellar Pocket ◽  
Novel Technologies ◽  
Near Future

Trypanosoma brucei is one of only a few unicellular pathogens that thrives extracellularly in the vertebrate host. Consequently, the cell surface plays a critical role in both immune recognition and immune evasion. The variant surface glycoprotein (VSG) coats the entire surface of the parasite and acts as a flexible shield to protect invariant proteins against immune recognition. Antigenic variation of the VSG coat is the major virulence mechanism of trypanosomes. In addition, incessant motility of the parasite contributes to its immune evasion, as the resulting fluid flow on the cell surface drags immunocomplexes toward the flagellar pocket, where they are internalized. The flagellar pocket is the sole site of endo- and exocytosis in this organism. After internalization, VSG is rapidly recycled back to the surface, whereas host antibodies are thought to be transported to the lysosome for degradation. For this essential step to work, effective machineries for both sorting and recycling of VSGs must have evolved in trypanosomes. Our understanding of the mechanisms behind VSG recycling and VSG secretion, is by far not complete. This review provides an overview of the trypanosome secretory and endosomal pathways. Longstanding questions are pinpointed that, with the advent of novel technologies, might be answered in the near future.

scholarly journals Endocytosis of a Glycosylphosphatidylinositol-anchored Protein via Clathrin-coated Vesicles, Sorting by Default in Endosomes, and Exocytosis via RAB11-positive Carriers

2003 ◽  
Vol 14 (5) ◽  
pp. 2029-2040 ◽  
Author(s):  
Christoph G. Grünfelder ◽  
Markus Engstler ◽  
Frank Weise ◽  
Heinz Schwarz ◽  
York-Dieter Stierhof ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Cell Surface ◽  
Trypanosoma Brucei ◽  
Coated Vesicles ◽  
Surface Glycoprotein ◽  
Variant Surface Glycoprotein ◽  
Flagellar Pocket ◽  
Protozoan Parasites ◽  
Basic Features ◽  
Membrane Concentration

Recently, proteins linked to glycosylphosphatidylinositol (GPI) residues have received considerable attention both for their association with lipid microdomains and for their specific transport between cellular membranes. Basic features of trafficking of GPI-anchored proteins or glycolipids may be explored in flagellated protozoan parasites, which offer the advantage that their surface is dominated by these components. In Trypanosoma brucei, the GPI-anchored variant surface glycoprotein (VSG) is efficiently sorted at multiple intracellular levels, leading to a 50-fold higher membrane concentration at the cell surface compared with the endoplasmic reticulum. We have studied the membrane and VSG flow at an invagination of the plasma membrane, the flagellar pocket, the sole region for endo- and exocytosis in this organism. VSG enters trypanosomes in large clathrin-coated vesicles (135 nm in diameter), which deliver their cargo to endosomes. In the lumen of cisternal endosomes, VSG is concentrated by default, because a distinct class of small clathrin-coated vesicles (50–60 nm in diameter) budding from the cisternae is depleted in VSG. TbRAB11-positive cisternal endosomes, containing VSG, fragment by an unknown process giving rise to intensely TbRAB11- as well as VSG-positive, disk-like carriers (154 nm in diameter, 34 nm in thickness), which are shown to fuse with the flagellar pocket membrane, thereby recycling VSG back to the cell surface.

African trypanosomes: the genome and adaptations for immune evasion

2011 ◽  
Vol 51 ◽  
pp. 47-62 ◽  
Author(s):  
Gloria Rudenko
Keyword(s):  
Immune Evasion ◽  
Cell Biology ◽  
Antigenic Variation ◽  
Critical Role ◽  
Sub Saharan Africa ◽  
Surface Glycoprotein ◽  
Tsetse Flies ◽  
African Trypanosomes ◽  
Genes Encoding ◽  
Sub Saharan

The African trypanosome Trypanosoma brucei is a flagellated unicellular parasite transmitted by tsetse flies that causes African sleeping sickness in sub-Saharan Africa. Trypanosomes are highly adapted for life in the hostile environment of the mammalian bloodstream, and have various adaptations to their cell biology that facilitate immune evasion. These include a specialized morphology, with most nutrient uptake occurring in the privileged location of the flagellar pocket. In addition, trypanosomes show extremely high rates of recycling of a protective VSG (variant surface glycoprotein) coat, whereby host antibodies are stripped off of the VSG before it is re-used. VSG recycling therefore functions as a mechanism for cleaning the VSG coat, allowing trypanosomes to survive in low titres of anti-VSG antibodies. Lastly, T. brucei has developed an extremely sophisticated strategy of antigenic variation of its VSG coat allowing it to evade host antibodies. A single trypanosome has more than 1500 VSG genes, most of which are located in extensive silent arrays. Strikingly, most of these silent VSGs are pseudogenes, and we are still in the process of trying to understand how non-intact VSGs are recombined to produce genes encoding functional coats. Only one VSG is expressed at a time from one of approximately 15 telomeric VSG ES (expression site) transcription units. It is becoming increasingly clear that chromatin remodelling must play a critical role in ES control. Hopefully, a better understanding of these unique trypanosome adaptations will eventually allow us to disrupt their ability to multiply in the mammalian bloodstream.

scholarly journals Trypanosoma brucei ribonuclease H2A is an essential enzyme that resolves R-loops associated with transcription initiation and antigenic variation

2019 ◽  
Author(s):  
Emma Briggs ◽  
Kathryn Crouch ◽  
Leandro Lemgruber ◽  
Graham Hamilton ◽  
Craig Lapsley ◽  
...  
Keyword(s):  
Gene Expression ◽  
Trypanosoma Brucei ◽  
Transcription Initiation ◽  
Antigenic Variation ◽  
Critical Role ◽  
Rnase H ◽  
Surface Glycoprotein ◽  
Pol Ii ◽  
Pol I ◽  
Rnase H2

In every cell ribonucleotides represent a threat to the stability and transmission of the DNA genome. Two types of Ribonuclease H (RNase H) tackle such ribonucleotides, either by excision when they form part of the DNA strand, or by hydrolysing RNA when it base-pairs with DNA, in structures termed R-loops. Loss of either RNase H is lethal in mammals, whereas yeast can prosper in the absence of both enzymes. Removal of RNase H1 is tolerated by the parasite Trypanosoma brucei but no work has examined the function of RNase H2. Here we show that loss of the catalytic subunit of T. brucei RNase H2 (TbRH2A) leads to growth and cell cycle arrest that is concomitant with accumulation of nuclear damage at sites of RNA polymerase (Pol) II transcription initiation, revealing a novel and critical role for RNase H2. In addition, differential gene expression of both RNA Pol I and II transcribed genes occurs after TbRH2A loss, including patterns that may relate to cytosolic DNA accumulation in humans with autoimmune disease. Finally, we show that TbRH2A loss causes R-loop and DNA damage accumulation in telomeric RNA Pol I transcription sites, leading to altered variant surface glycoprotein expression. Thus, we demonstrate a separation of function between the two nuclear T. brucei RNase H enzymes during RNA Pol II transcription, but overlap in function during RNA Pol I-mediated gene expression during host immune evasion.

scholarly journals Black-necked spitting cobra (Naja nigricollis) phospholipases A2 cause Trypanosoma brucei death by blocking endocytosis through the flagellar pocket

2021 ◽  
Author(s):  
Andrea Martos-Esteban ◽  
Olivia J. S. Macleod ◽  
Isabella Maudlin ◽  
Konstantinos Kalogeropoulos ◽  
Jonas A. Jurgensen ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Trypanosoma Brucei ◽  
Antigenic Variation ◽  
Adaptive Immune Response ◽  
Surface Glycoprotein ◽  
Covalent Attachment ◽  
Mammalian Host ◽  
Flagellar Pocket ◽  
African Trypanosomes ◽  
External Face

African trypanosomes, such as Trypanosoma brucei, are flagellated protozoa which proliferate in mammals and cause a variety of diseases in people and animals. In a mammalian host, the external face of the African trypanosome plasma membrane is covered by a densely packed coat formed of variant surface glycoprotein (VSG), which counteracts the host adaptive immune response by antigenic variation. The VSG is attached to the external face of the plasma membrane by covalent attachment of the C-terminus to a glycosylphosphatidylinositol. As the trypanosome grows, newly synthesised VSG is added to the plasma membrane by vesicle fusion to the flagellar pocket, the sole location of exo- and endocytosis. Snake venoms contain dozens of components including proteases and phospholipases. Here, we investigated the effect of Naja nigricollis on T. brucei with the aim of describing the response of the trypanosome to hydrolytic attack on the VSG. We found no evidence for VGS hydrolysis however N. nigricollis venom caused: (i) an enlargement of the flagellar pocket, (ii) the Rab11 positive endosomal compartments to adopt an abnormal dispersed localisation, and (iii) a cell cycle arrest prior to cytokinesis. A single protein family, the phospholipases A2s present in N. nigricollis venom, was necessary and sufficient for the effects. This study provides new molecular insight into T. brucei biology and possibly describes mechanisms that could be exploited for T. brucei targeting.

scholarly journals Characterization of a cDNA encoding a cysteine-rich cell surface protein located in the flagellar pocket of the protozoan Trypanosoma brucei.

1990 ◽  
Vol 10 (9) ◽  
pp. 4506-4517 ◽  
Author(s):  
M G Lee ◽  
B E Bihain ◽  
D G Russell ◽  
R J Deckelbaum ◽  
L H Van der Ploeg
Keyword(s):  
Amino Acid ◽  
Cell Surface ◽  
Trypanosoma Brucei ◽  
Transmembrane Domain ◽  
Density Lipoprotein ◽  
Surface Protein ◽  
Integral Membrane Protein ◽  
Cell Surface Receptor ◽  
Flagellar Pocket ◽  
Cytoplasmic Extension

We have characterized a cDNA encoding a cysteine-rich, acidic integral membrane protein (CRAM) of the parasitic protozoa Trypanosoma brucei and Trypanosoma equiperdum. Unlike other membrane proteins of T. brucei, which are distributed throughout the cell surface, CRAM is concentrated in the flagellar pocket, an invagination of the cell surface of the trypanosome where endocytosis has been documented. Accordingly, CRAM also locates to vesicles located underneath the pocket, providing evidence of its internalization. CRAM has a predicted molecular mass of 130 kilodaltons and has a signal peptide, a transmembrane domain, and a 41-amino-acid cytoplasmic extension. A characteristic feature of CRAM is a large extracellular domain with a roughly 66-fold acidic, cysteine-rich 12-amino-acid repeat. CRAM is conserved among different protozoan species, including Trypanosoma cruzi, and CRAM has structural similarities with eucaryotic cell surface receptors. The most striking homology of CRAM is to the human low-density-lipoprotein receptor. We propose that CRAM functions as a cell surface receptor of different trypanosome species.

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 Chromosome structure: DNA nucleotide sequence elements of a subset of the minichromosomes of the protozoan Trypanosoma brucei.

1991 ◽  
Vol 11 (8) ◽  
pp. 3823-3834 ◽  
Author(s):  
M Weiden ◽  
Y N Osheim ◽  
A L Beyer ◽  
L H Van der Ploeg
Keyword(s):  
Trypanosoma Brucei ◽  
Antigenic Variation ◽  
Microscopic Analysis ◽  
The Body ◽  
Surface Glycoprotein ◽  
Main Body ◽  
Electron Microscopic ◽  
Protein Coding ◽  
Cell Surface Glycoprotein ◽  
Telomere Repeat

The genome of the protozoan Trypanosoma brucei contains a set of about 100 minichromosomes of about 50 to 150 kb in size. The small size of these chromosomes, their involvement in antigenic variation, and their mitotic stability make them ideal candidates for a structural analysis of protozoan chromosomes and their telomeres. We show that a subset of the minichromosomes is composed predominantly of simple-sequence DNA, with over 90% of the length of the minichromosome consisting of a tandem array of 177-bp repeats, indicating that these molecules have limited protein-coding capacity. Proceeding from the tip of the telomere to a chromosome internal position, a subset of the minichromosomes contained the GGGTTA telomere repeat, a 29-bp telomere-derived repeat, a region containing 74-bp G + C-rich direct repeats separated by approximately 155 bp of A + T-rich DNA that has a bent character, and 50 to 150 kb of the 177-bp repeat. Several of the minichromosome-derived telomeres did not encode protein-coding genes, indicating that the repertoire of telomeric variant cell surface glycoprotein genes is restricted to some telomeres only. The telomere organization in trypanosomes shares striking similarities to the organization of telomeres and subtelomeres in humans, yeasts, and plasmodia. An electron microscopic analysis of the minichromosomes showed that they are linear molecules without abnormal structures in the main body of the chromosome. The structure of replicating molecules indicated that minichromosomes probably have a single bidirectional origin of replication located in the body of the chromosome. We propose a model for the structure of the trypanosome minichromosomes.

scholarly journals Trypanosoma brucei PolIE suppresses telomere recombination

2021 ◽  
Author(s):  
Maiko Tonini ◽  
M. A. G. Rabbani ◽  
Marjia Afrin ◽  
Bibo Li
Keyword(s):  
Trypanosoma Brucei ◽  
Immune Evasion ◽  
Human African Trypanosomiasis ◽  
Antigenic Variation ◽  
Telomere Maintenance ◽  
Genome Integrity ◽  
Major Player ◽  
Elevated Rate ◽  
Major Surface ◽  
Telomere Structure

Telomeres are essential for genome integrity and stability. In T. brucei that causes human African trypanosomiasis, the telomere structure and telomere proteins also influence the virulence of the parasite, as its major surface antigen involved in the host immune evasion is expressed exclusively from loci immediately upstream of the telomere repeats. However, telomere maintenance mechanisms are still unclear except that telomerase-mediated telomere synthesis is a major player. We now identify PolIE as an intrinsic telomere complex component. We find that depletion of PolIE leads to an increased amount of telomere/subtelomere DNA damage, an elevated rate of antigenic variation, and an increased amount of telomere T-circles and C-circles, indicating that PolIE suppresses telomere recombination and helps maintain telomere integrity. In addition, we observe much longer telomere G-rich 3 prime overhangs in PolIE-depleted cells, which is not dependent on telomerase. Furthermore, the level of telomere DNA synthesis is slightly increased in PolIE-depleted cells, which is dependent on telomerase. Therefore, we identify PolIE as a major player for telomere maintenance in T. brucei.

scholarly journals RibonucleaseH1-targeted R-loops in surface antigen gene expression sites can direct trypanosome immune evasion

2018 ◽  
Author(s):  
Emma Briggs ◽  
Kathryn Crouch ◽  
Leandro Lemgruber ◽  
Craig Lapsley ◽  
Richard McCulloch
Keyword(s):  
Immune Evasion ◽  
Protein Gene ◽  
Antigenic Variation ◽  
Surface Protein ◽  
Surface Glycoprotein ◽  
Surface Coat ◽  
Loop Formation ◽  
New Host ◽  
Genome Damage ◽  
Surface Protein Gene

AbstractSwitching of the Variant Surface Glycoprotein (VSG) inTrypanosoma bruceiprovides a crucial host immune evasion strategy that is catalysed both by transcription and recombination reactions, each operating within specialised telomeric VSG expression sites (ES). VSG switching is likely triggered by events focused on the single actively transcribed ES, from a repertoire of around 15, but the nature of such events is unclear. Here we show that RNA-DNA hybrids, called R-loops, form preferentially within sequences termed the 70 bp repeats in the actively transcribed ES, but spread throughout the active and inactive ES in the absence of RNase H1, which degrades R-loops. Loss of RNase H1 also leads to increased levels of VSG coat switching and replication-associated genome damage, some of which accumulates within the active ES. This work indicates VSG ES architecture elicits R-loop formation, and that these RNA-DNA hybrids connectT. bruceiimmune evasion by transcription and recombination.Author summaryAll pathogens must survive eradication by the host immune response in order to continue infections and be passed on to a new host. Changes in the proteins expressed on the surface of the pathogen, or on the surface of the cells the pathogen infects, is a widely used strategy to escape immune elimination. Understanding how this survival strategy, termed antigenic variation, operates in any pathogen is critical, both to understand interaction between the pathogen and host and disease progression. A key event in antigenic variation is the initiation of the change in expression of the surface protein gene, though how this occurs has been detailed in very few pathogens. Here we examine how changes in expression of the surface coat of the African trypanosome, which causes sleeping sickness disease, are initiated. We reveal that specialised nucleic acid structures, termed R-loops, form around the expressed trypanosome surface protein gene and increase in abundance after mutation of an enzyme that removes them, leading to increased changes in the surface coat in trypanosome cells that are dividing. We therefore shed light on the earliest acting events in trypanosome antigenic variation.

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