Aneuploidy is widely observed in both unicellular and multicellular eukaryotes, usually associated with adaptation to stress conditions. Chromosomal duplication stability a tradeoff between the fitness cost of having unbalanced gene copies potential gained from increased dosage specific advantageous genes. Trypanosomatids, family protozoans that include species cause neglected tropical diseases, are relevant group study aneuploidies. Their life cycle has several stressors could select for...

DNA replication initiates on defined genome sites, termed origins. Origin usage appears to follow common rules in the eukaryotic organisms examined date: all chromosomes are replicated from multiple origins, which display variations firing efficiency and selected a larger pool of potential To ask if these features true eukaryotes, we describe genome-wide origin mapping parasite Leishmania. Leishmania suggests striking divergence relative characterized since each chromosome be single origin....

Survival of Trypanosoma brucei depends upon switches in its protective Variant Surface Glycoprotein (VSG) coat by antigenic variation. VSG switching occurs frequent homologous recombination, which is thought to require locus-specific initiation. Here, we show that a RecQ helicase, RECQ2, acts repair DNA breaks, including the telomeric site expression. Despite this, RECQ2 loss does not impair variation, but causes increased arguing against models for switch initiation through direct...

African trypanosomes proliferate as bloodstream forms (BSFs) and procyclic in the mammal tsetse fly midgut, respectively. This allows them to colonise host environment upon infection ensure life cycle progression. Yet, understanding of mechanisms that regulate drive cell replication these is limited. Using single-cell transcriptomics on unsynchronised populations, we have obtained high resolution regulated (CCR) transcriptomes both slender BSF Trypanosoma brucei without prior sorting or...

RNA-DNA hybrids are epigenetic features of all genomes that intersect with many processes, including transcription, telomere homeostasis, and centromere function. Increasing evidence suggests can provide two conflicting roles in the maintenance transmission genomes: They be triggers DNA damage, leading to genome change, or aid repair processes needed respond lesions. Evasion host immunity by African trypanosomes, such as Trypanosoma brucei, relies on targeted recombination silent Variant...

Initiation of DNA replication depends upon recognition genomic sites, termed origins, by AAA+ ATPases. In prokaryotes a single factor binds each origin, whereas in eukaryotes this role is played six-protein origin complex (ORC). Why evolved multisubunit initiator, and the roles component, remains unclear. Trypanosoma brucei, an ancient unicellular eukaryote, only one ORC-related TbORC1/CDC6, has been identified sequence homology. Here we show that three TbORC1/CDC6-interacting factors also...

Chromosome damage must be repaired to prevent the proliferation of defective cells. Alternatively, cells with eliminated. This is true human and several other cell types but may not case for single-celled parasites, such as trypanosomes. African trypanosomes, which cause lethal diseases in both humans livestock, can actually exploit chromosomal activate new surface coat proteins evade host immune responses, example. We monitored responses single breaks trypanosomes using a DNA-binding...

The Trypanosoma brucei genome is structurally complex. Eleven megabase-sized chromosomes each comprise a transcribed core flanked by silent subtelomeres, housing thousands of Variant Surface Glycoprotein (VSG) genes. Additionally, hundreds sub-megabase contain 177 bp repeats unknown function, and VSG transcription sites localise to many telomeres. DNA replication dynamics have only been described in the megabase chromosome cores, single active site. Using Nanopore assembly, we show that...

Abstract Genomes in eukaryotes normally undergo DNA replication a choreographed temporal order, resulting early and late replicating chromosome compartments. Leishmania , human protozoan parasite, displays an unconventional program which the timing of completion is size-dependent: larger chromosomes complete later then smaller ones. Here we show that both R-loops RNase H1, ribonuclease resolves RNA-DNA hybrids, accumulate major pattern reflects their timing. Furthermore, demonstrate such...

Bloodstream-form African trypanosomes display mono-telomeric expression of a Variant Surface Glycoprotein (VSG) gene in an inter-chromosomally bridged transcription and splicing compartment, such that the dominant produces 10,000 times more transcript than excluded VSG genes. Antigenic variation, whereby parasites switch to express other VSGs, then underpins robust host immune evasion strategy. Specific chromatin RNA-associated factors are required maintain exclusion, but our understanding...

Peptidomimetic imidazolidin-4-one derivatives of primaquine (imidazoquines) recently displayed in vitro activity against blood schizonts a chloroquine-resistant strain Plasmodium falciparum. Preliminary studies with subset such imidazoquines showed them to both block transmission P. berghei malaria from mouse mosquito and be highly stable toward hydrolysis at physiological conditions. This prompted us have deeper insight into the Plasmodia Pneumocystis carinii, on which is also active. Full...

Eukaryotic genome duplication relies on origins of replication, distributed over multiple chromosomes, to initiate DNA replication. A recent genome-wide analysis Trypanosoma brucei, the etiological agent sleeping sickness, localized its replication boundaries multigenic transcription units. To better understand genomic in this organism, we examined by single molecule replicated DNA. We determined average speed forks procyclic and bloodstream form cells found that T. brucei rate is similar...

DNA replication is needed to duplicate a cell’s genome in S phase and segregate it during cell division. Previous work Leishmania detected initiation at just single region each chromosome, an organisation predicted be insufficient for complete duplication within phase. Here, we show that acetylated histone H3 (AcH3), base J kinetochore factor co-localise chromosome only locus, which corresponds with previously mapped regions demarcated by localised G/T skew G4 patterns. In addition, describe...

Abstract The genome of Trypanosoma brucei is structurally complex. Eleven megabase-sized chromosomes each comprise a transcribed core flanked by silent subtelomeres, housing thousands Variant Surface Glycoprotein ( VSG ) genes. Additionally, VSGs are also found on hundreds sub-megabase that harbour 177 bp repeats unknown function, and multiple transcription sites localise to the telomeres both chromosome types. DNA replication dynamics have been described in megabase cores but not...

Summary Nucleotide excision repair ( NER ) is a highly conserved genome pathway acting on helix distorting DNA lesions. divided into two subpathways: global GG‐NER ), which responsible for throughout genomes, and transcription‐coupled TC‐NER acts lesions that impede transcription. The extent of the T rypanosoma brucei transcribed unusual, since most genes are organized in multigene transcription units, each from single promoter. Given this organization, we have addressed importance to ....

Abstract Trypanosomatids, which include major pathogens of humans and livestock, are flagellated protozoa for cell cycle controls the underlying mechanisms not completely understood. Here, we describe a genome-wide RNA-interference library screen defects in Trypanosoma brucei . We induced massive parallel knockdown, sorted perturbed population using high-throughput flow cytometry, deep-sequenced RNAi-targets from each stage digitally reconstructed profiles at genomic scale; also enabling...

Abstract Background DNA replication in trypanosomatids operates a uniquely challenging environment, since most of their genomes are constitutively transcribed. Trypanosoma cruzi , the etiological agent Chagas disease, presents high variability both chromosomes size and copy number among strains, though underlying mechanisms unknown. Results Here we have mapped sites initiation across T. genome using Marker Frequency Analysis, which has previously only been deployed two related...

Abstract Genomes in eukaryotes normally undergo DNA replication a choreographed temporal order, resulting early and late replicating chromosome compartments. Leishmania , human protozoan parasite, displays an unconventional program which the timing of completion is size-dependent: larger chromosomes complete later then smaller ones. Here we show that both R-loops RNase H1, ribonuclease resolves RNA-DNA hybrids, accumulate major pattern reflects their timing. Furthermore, demonstrate such...

Abstract RNA–DNA hybrids are epigenetic features of genomes that provide a diverse and growing range activities. Understanding these functions has been informed by characterising the proteins interact with hybrids, but all such analyses have so far focused on mammals, meaning it is unclear if similar spectrum hybrid interactors found in other eukaryotes. The African trypanosome single-cell eukaryotic parasite Discoba grouping displays substantial divergence several aspects core biology from...

Parasite infection can lead to alterations in the permeability of host plasma membranes. Presented here is first demonstration that this phenomenon occurs Plasmodium-infected liver cells. Using whole-cell patch-clamp technique, volume-regulated anion channel (VRAC) activity was characterized Huh-7 cells (a human hepatoma cell line) before and after with Plasmodium berghei. Consistent presence VRACs, hypotonic bath solution induced large ion currents rectified outwardly, reversed close...