Control of Variant Surface Glycoprotein Expression by CFB2 in Trypanosoma brucei and Quantitative Proteomic Connections to Translation and Cytokinesis

Ruiz, Gustavo Bravo; Tinti, Michele; Ridgway, Melanie C.; Horn, David

doi:10.1128/msphere.00069-22

Cited by 6 publications

(3 citation statements)

References 43 publications

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“…Consistent with this, these cells were marked by relatively high expression, for example, of EP and GPEET procyclins, ZC3H20, PAD2, and ZFPs, each of which has been found to be up-regulated as parasites prepare for transmission or undergo stumpy formation ( 10 , 23 ). The cyclin F box2 protein was also elevated, this being associated with VSG mRNA stability ( 24 ).…”

Section: Resultsmentioning

confidence: 99%

The developmental hierarchy and scarcity of replicative slender trypanosomes in blood challenges their role in infection maintenance

Larcombe,

Briggs,

Savill

et al. 2023

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

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The development of Trypanosoma brucei in its mammalian host is marked by a distinct morphological change as replicative “slender” forms differentiate into cell cycle arrested “stumpy” forms in a quorum-sensing-dependent manner. Although stumpy forms dominate chronic infections at the population level, the proportion of replicative parasites at the individual cell level and the irreversibility of arrest in the bloodstream are unclear. Here, we experimentally demonstrate that developmental cell cycle arrest is definitively irreversible in acute and chronic infections in mice. Furthermore, analysis of replicative capacity and single-cell transcriptome profiling reveal a temporal hierarchy, whereby cell cycle arrest and appearance of a reversible stumpy-like transcriptome precede irreversible commitment and morphological change. Unexpectedly, we show that proliferating parasites are exceptionally scarce in the blood after infections are established. This challenges the ability of bloodstream trypanosomes to sustain infection by proliferation or antigenic variation, these parasites instead being overwhelmingly adapted for transmission.

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Section: Resultsmentioning

confidence: 99%

The developmental hierarchy and scarcity of replicative slender trypanosomes in blood challenges their role in infection maintenance

Larcombe,

Briggs,

Savill

et al. 2023

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

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“…One of these mechanisms proposes the presence of a 16mer binding factor which prevents degradation of the mRNA by blocking association of the transcript with nucleases (Ridewood et al ., 2017). CFB2 has been identified as the 16mer binding factor and further shown to act by recruiting a stabilising complex that includes MKT1, PBP1, PABP2, and the cap-binding translation initiation complex EIF4E6/G5 (do Nascimento et al ., 2021; Bravo Ruiz et al ., 2022). The second proposed mechanism involves a 16mer dependent m 6 A modification of poly(A) tails of VSG transcripts which inhibits RNA degradation by blocking deadenylation (Viegas et al ., 2022).…”

Section: Discussionmentioning

confidence: 99%

The dual role of the 16mer motif within the 3’ untranslated region of the variant surface glycoprotein ofTrypanosoma brucei

Bakari-Soale,

Batram,

Zimmerman

et al. 2023

Preprint

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The variant surface glycoprotein (VSG) of African trypanosomes is essential for survival of bloodstream form parasites. These parasites undergo antigenic variation, an immune evasion strategy in which they periodically switch VSG expression from one isoform to another. The molecular processes central to the expression and regulation of the VSG are however not fully understood. In general, the regulation of gene expression in trypanosomes is largely post-transcriptional. Regulatory sequences, mostly present in the 3’ UTRs, often serve as key elements in the modulation of the levels of individual mRNAs. InT. bruceiVSG genes, a 16mer motif within the 3’ UTR has been shown to be essential for the stability ofVSGtranscripts and abundant VSG expression. This motif is 100 % conserved in the 3’ UTRs of all transcribed and non-transcribed VSG genes. As a stability-associated sequence element, the absence of nucleotide substitutions in the 16mer is however exceptional. We therefore hypothesised that the motif is involved in other essential roles/processes besides stability of theVSGtranscripts.In this study, we demonstrate that the 100 % conservation of the 16mer motif is not essential for cell viability or for the maintenance of functional VSG protein levels. We further show that the intact motif in the active VSG 3’ UTR is neither required to promote VSG silencing during switching nor is it needed during differentiation from bloodstream forms to procyclic forms. Ectopic overexpression of a second VSG, however, requires the intact 16mer motif within the ectopic VSG 3’ UTR to trigger silencing and exchange of the active VSG, suggesting a role for the motif in transcriptional VSG switching. The enigmatic 16mer motif therefore appears to play a dual role in transcriptionalVSGswitching andVSGtranscript stability.

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“…In addition, the total amount of VSG mRNA appears to be fixed, as expressing an ectopic VSG gene leads to downregulation of the endogenous VSG expression [286,[288][289][290][291]. This is possibly regulated through the 16-mer VSG 3 ′ UTR consensus sequence [288], which can be bound by CFB2 [292,293] and modified with m6A [294], both important for VSG mRNA stability. However, adding stop codons in the VSG gene ORF leads to dramatically increased total VSG mRNA levels, suggesting that translation of VSG mRNA is actually monitored to regulate VSG expression [295].…”

Section: Competition Between Tbrap1's Dna and Rna Binding Activities ...mentioning

confidence: 99%

Unwrap RAP1’s Mystery at Kinetoplastid Telomeres

2024

Biomolecules

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Although located at the chromosome end, telomeres are an essential chromosome component that helps maintain genome integrity and chromosome stability from protozoa to mammals. The role of telomere proteins in chromosome end protection is conserved, where they suppress various DNA damage response machineries and block nucleolytic degradation of the natural chromosome ends, although the detailed underlying mechanisms are not identical. In addition, the specialized telomere structure exerts a repressive epigenetic effect on expression of genes located at subtelomeres in a number of eukaryotic organisms. This so-called telomeric silencing also affects virulence of a number of microbial pathogens that undergo antigenic variation/phenotypic switching. Telomere proteins, particularly the RAP1 homologs, have been shown to be a key player for telomeric silencing. RAP1 homologs also suppress the expression of Telomere Repeat-containing RNA (TERRA), which is linked to their roles in telomere stability maintenance. The functions of RAP1s in suppressing telomere recombination are largely conserved from kinetoplastids to mammals. However, the underlying mechanisms of RAP1-mediated telomeric silencing have many species-specific features. In this review, I will focus on Trypanosoma brucei RAP1’s functions in suppressing telomeric/subtelomeric DNA recombination and in the regulation of monoallelic expression of subtelomere-located major surface antigen genes. Common and unique mechanisms will be compared among RAP1 homologs, and their implications will be discussed.

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Control of Variant Surface Glycoprotein Expression by CFB2 in Trypanosoma brucei and Quantitative Proteomic Connections to Translation and Cytokinesis

Abstract: VSG expression represents a key parasite virulence mechanism and an example of extreme biology. Posttranscriptional gene expression controls in trypanosomatids also continue to be the subject of substantial research interest.

Cited by 6 publications

References 43 publications

The developmental hierarchy and scarcity of replicative slender trypanosomes in blood challenges their role in infection maintenance

The developmental hierarchy and scarcity of replicative slender trypanosomes in blood challenges their role in infection maintenance

The dual role of the 16mer motif within the 3’ untranslated region of the variant surface glycoprotein ofTrypanosoma brucei

Unwrap RAP1’s Mystery at Kinetoplastid Telomeres

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