Suppression of subtelomeric VSG switching by Trypanosoma brucei TRF requires its TTAGGG repeat-binding activity

Jehi, Sanaa E.; Li, Xiaohua; Sandhu, Ranjodh; Ye, Fei; Benmerzouga, Imaan; Zhang, Mingjie; Zhao, Yanxiang; Li, Bibo

doi:10.1093/nar/gku942

Cited by 44 publications

(89 citation statements)

References 75 publications

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“…These findings indicate that the telomeric NPT reporter is subject to the exclusion system operating in T. brucei. Analysis of Tb927.6.4430 supported previous reports of disrupted VSG silencing when telomere structure and/or function are compromised (26,34); knockdown of this novel telomeric TTAGGG repeat-associated factor (SI Appendix, Fig. S2 A and B) was associated with reporter and VSG derepression (SI Appendix, Fig.…”

Section: Significancesupporting

confidence: 80%

“…2C and Movie S2) that is the site of the single active VSG-ES (10). Because all competent VSG-ESs are telomeric, we also used superresolution microscopy to assess VEX1 localization relative to the telomeric TTAGGG repeat-binding factor TRF2 (34). This revealed punctate nuclear TRF2 staining and VEX1 staining that was coincident with a TRF2 punctum (Fig.…”

Section: Significancementioning

confidence: 99%

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VEX1 controls the allelic exclusion required for antigenic variation in trypanosomes

Glover

Hutchinson

Alsford

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

184

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Allelic exclusion underpins antigenic variation and immune evasion in African trypanosomes. These bloodstream parasites use RNA polymerase-I (pol-I) to transcribe just one telomeric variant surface glycoprotein (VSG) gene at a time, producing superabundant and switchable VSG coats. We identified trypanosome VSG exclusion-1 (VEX1) using a genetic screen for defects in telomere-exclusive expression. VEX1 was sequestered by the active VSG and silencing of other VSGs failed when VEX1 was either ectopically expressed or depleted, indicating positive and negative regulation, respectively. Positive regulation affected VSGs and nontelomeric pol-I-transcribed genes, whereas negative regulation primarily affected VSGs. Negative regulation by VEX1 also affected telomeric pol-I-transcribed reporter constructs, but only when they contained blocks of sequence sharing homology with a pol-I-transcribed locus. We conclude that restricted positive regulation due to VEX1 sequestration, combined with VEX1-dependent, possibly homology-dependent silencing, drives a "winner-takes-all" mechanism of allelic exclusion.epigenetic | monoallelic | silencing | telomere | Trypanosoma brucei C ells often restrict expression to a single allele of a gene or gene family. This allelic exclusion underpins antigenic variation in pathogens, including trypanosomes that cause sleeping sickness (1) and Plasmodium parasites that cause malaria (2). Allelic exclusion is also essential for singular olfactory receptor expression and a sense of smell in metazoa (3). Although many factors have been identified that are required for the expression of one allele or for the silencing of other alleles in these systems, our understanding of the mechanisms by which expression and silencing are established and coordinated remains incomplete (1-3).The African trypanosome Trypanosoma brucei is a flagellated parasitic protozoan transmitted among mammalian hosts by tsetse flies. In addition to causing trypanosomiasis in humans, a fatal and neglected tropical disease, these parasites also cause nagana in cattle. Antigenic variation is essential for persistent bloodstream infection in the face of host adaptive immune defenses and has long been a paradigm for studies on allelic exclusion (1); parasite immune evasion depends upon singular variant surface glycoprotein (VSG) gene expression and VSG switching. Although multiple subtelomeric VSGs are available for expression (4), only one is transcribed (5). Both active and silent VSGs are located at the ends of polycistronic transcription units known as expression sites (ESs) (6). Notably, VSG-ES promoters (6) recruit RNA polymerase-I (pol-I) that typically transcribes ribosomal RNA genes (7). Indeed, the active VSG-ES is associated with an extranucleolar focus of pol-I known as the expression-site body (ESB) (8-10). Although the active VSG-ES is specifically depleted of nucleosomes (11, 12), silent VSG-ESs are similarly located in the extranucleolar space in bloodstream-form cells, and neither active nor silent VSG-ESs show an ...

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Section: Significancesupporting

confidence: 80%

Section: Significancementioning

confidence: 99%

VEX1 controls the allelic exclusion required for antigenic variation in trypanosomes

Glover

Hutchinson

Alsford

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

184

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“…Active transcription is known to favor homologous recombination (37). RNAi knockdown of the telomeric proteins TbTRF or TbTRF-interacting factor 2 (TbTIF2) increased VSG switching rates in T. brucei BF (38,39). The loss of these proteins appears to have enhanced repair and recombination.…”

Section: Discussionmentioning

confidence: 99%

Inositol phosphate pathway controls transcription of telomeric expression sites in trypanosomes

Cestari

Stuart

2015

Proc. Natl. Acad. Sci. U.S.A.

143

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African trypanosomes evade clearance by host antibodies by periodically changing their variant surface glycoprotein (VSG) coat. They transcribe only one VSG gene at a time from 1 of about 20 telomeric expression sites (ESs). They undergo antigenic variation by switching transcription between telomeric ESs or by recombination of the VSG gene expressed. We show that the inositol phosphate (IP) pathway controls transcription of telomeric ESs and VSG antigenic switching in Trypanosoma brucei. Conditional knockdown of phosphatidylinositol 5-kinase (TbPIP5K) or phosphatidylinositol 5-phosphatase (TbPIP5-Pase) or overexpression of phospholipase C (TbPLC) derepresses numerous silent ESs in T. brucei bloodstream forms. The derepression is specific to telomeric ESs, and it coincides with an increase in the number of colocalizing telomeric and RNA polymerase I foci in the nucleus. Monoallelic VSG transcription resumes after reexpression of TbPIP5K; however, most of the resultant cells switched the VSG gene expressed. TbPIP5K, TbPLC, their substrates, and products localize to the plasma membrane, whereas TbPIP5Pase localizes to the nucleus proximal to telomeres. TbPIP5Pase associates with repressor/activator protein 1 (TbRAP1), and their telomeric silencing function is altered by TbPIP5K knockdown. These results show that specific steps in the IP pathway control ES transcription and antigenic switching in T. brucei by epigenetic regulation of telomere silencing.O nly one of the ∼20 telomeric expression sites (ESs) is transcribed at a time in Trypanosoma brucei in the mammalian infectious stage bloodstream (BF) or metacyclic forms (MFs), whereas no ES is transcribed in the insect stage procyclic forms (PFs) (1). Each ES contains 1 telomeric variant surface glycoprotein (VSG) gene, whose expression confers a distinct cellular antigenic type and up to 12 expression site-associated genes (ESAGs) whose functions are incompletely understood ( Fig. 1A) (2-4). The parasites evade immune clearance by periodically changing antigenic type by switching transcription between ESs or by ES recombination with the ∼2,500 non-ES VSG genes and pseudogenes (5, 6). ESs are transcribed by RNA polymerase I (Pol I), which initiates at the single promoter at all ESs but terminates within a few kilobases except at one fully transcribed ES (7,8).RNAi knockdown of expression of the nuclear protein TbRAP1 that interacts with the TTAGGG-binding factor (TbTRF) or of the nuclear lamina protein 1 (TbNUP1) or deletion of the histonelysine N-methyltransferase DOT1B (TbDOT1B) alleles each results in transcription of silent ESs (9-11). Similarly, RNAi knockdown of the transcription facilitating histone chaperone suppressor of ty 16 (TbSPT16) or the SWI2/SNF2-related chromatin-remodeling protein TbISWI or deletion of histone deacetylase sirtuin 2-related protein 1 (TbSIR2RP1) each increases transcription near the promoter but not the VSG gene of silent . These results indicate that the control of ES transcription involves telomeric silencing (15), which entails...

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“…Recently, we showed that telomere proteins are also important for maintaining subtelomere integrity and stability (24,25,27). In addition, telomeres suppress expression of nearby genes by telomeric silencing (28).…”

Section: Introductionmentioning

confidence: 99%

Trypanosoma brucei RAP1 maintains telomere and subtelomere integrity by suppressing TERRA and telomeric RNA:DNA hybrids

Nanavaty

Sandhu

Jehi

et al. 2017

Nucleic Acids Research

Self Cite

121

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Trypanosoma brucei causes human African trypanosomiasis and regularly switches its major surface antigen, VSG, thereby evading the host's immune response. VSGs are monoallelically expressed from subtelomeric expression sites (ESs), and VSG switching exploits subtelomere plasticity. However, subtelomere integrity is essential for T. brucei viability. The telomeric transcript, TERRA, was detected in T. brucei previously. We now show that the active ES-adjacent telomere is transcribed. We find that TbRAP1, a telomere protein essential for VSG silencing, suppresses VSG gene conversion-mediated switching. Importantly, TbRAP1 depletion increases the TERRA level, which appears to result from longer read-through into the telomere downstream of the active ES. Depletion of TbRAP1 also results in more telomeric RNA:DNA hybrids and more double strand breaks (DSBs) at telomeres and subtelomeres. In TbRAP1-depleted cells, expression of excessive TbRNaseH1, which cleaves the RNA strand of the RNA:DNA hybrid, brought telomeric RNA:DNA hybrids, telomeric/subtelomeric DSBs and VSG switching frequency back to WT levels. Therefore, TbRAP1-regulated appropriate levels of TERRA and telomeric RNA:DNA hybrid are fundamental to subtelomere/telomere integrity. Our study revealed for the first time an important role of a long, non-coding RNA in antigenic variation and demonstrated a link between telomeric silencing and subtelomere/telomere integrity through TbRAP1-regulated telomere transcription.

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Suppression of subtelomeric VSG switching by Trypanosoma brucei TRF requires its TTAGGG repeat-binding activity

Cited by 44 publications

References 75 publications

VEX1 controls the allelic exclusion required for antigenic variation in trypanosomes

VEX1 controls the allelic exclusion required for antigenic variation in trypanosomes

Inositol phosphate pathway controls transcription of telomeric expression sites in trypanosomes

Trypanosoma brucei RAP1 maintains telomere and subtelomere integrity by suppressing TERRA and telomeric RNA:DNA hybrids

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