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

Larcombe, Stephen D.; Briggs, Emma M.; Savill, Nick; Szoor, Balazs; Matthews, Keith R.

doi:10.1073/pnas.2306848120

Cited by 8 publications

(30 citation statements)

References 47 publications

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“…In this study, we performed m 6 A immunoprecipitation without prior RNA fragmentation to obtain a complete set of methylated transcripts, regardless of the distribution of m 6 A modifications in poly(A) tails [16] or in internal locations [15]. Therefore, we did not assess which transcripts are internally or poly(A) methylated, and whether that impacts their expression and/or stability.…”

Section: Discussionmentioning

confidence: 99%

“…We also detected another RNA binding protein, a zinc-finger protein, a hypothetical protein, and a cyclinlike F-box 2 (CFB2) protein within the most abundant transcripts. Analysis of transcripts from procyclic forms showed again a silent VSG as the transcript most enriched in m 6 A, followed by a hypothetical protein, a zinc-finger protein, separase and one expression site-associated gene 2 (ESAG2) (Fig. 3D).…”

Section: Identification Of M 6 A-methylated Transcripts In Three Life...mentioning

confidence: 97%

“…The function of m 6 A varies between organisms, including parasites, where it might be important for life cycle progression. In Toxoplasma gondii, m 6 A methylation is prevalent in asexual life cycle stages [12] and is involved in proper mRNA 3' end processing, likely affecting developmental gene regulation [13]. In Plasmodium falciparum, m 6 A is highly developmentally regulated, mediating translational repression of transcripts involved in regulation of gene expression in blood stages [14].…”

Section: Introductionmentioning

confidence: 99%

“…Their study showed how both life cycle stages differed in terms of methylated transcripts and identified in which region of the transcript (coding or untranslated) m 6 A peaks were located [15]. More recently, our lab identified m 6 A can also be found in the poly(A) tail, where it stabilizes VSG transcripts in slender forms and whose removal preceded VSG downregulation during parasite differentiation to procyclic forms [16]. However, whether m 6 A methylation is important for gene expression regulation in other stages of the T. brucei life cycle remains unknown.…”

Section: Introductionmentioning

confidence: 99%

“…When slender forms reach a critical cell density in the bloodstream, they differentiate into non-replicative stumpy forms through a quorum-sensing mechanism triggered by oligopeptides, generally known as the stumpy induction factor (SIF). Such peptides are generated by parasite-released peptidases in a cell density-dependent manner and activate a signaling pathway that leads to gene expression remodeling, including reduction of transcription and translation [5][6][7][8]. Uptake of bloodstream parasites by the tsetse fly triggers further differentiation into procyclic forms, which lead to broad transcriptomic and morphological changes, including the replacement of the VSG by a procyclin surface coat [9].…”

Section: Introductionmentioning

confidence: 99%

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m⁶A landscape is more pervasive whenTrypanosoma bruceiexits the cell cycle

Serra,

Pereira,

Viegas

et al. 2024

Preprint

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N6-methyladenosine (m6A) is a mRNA modification with important roles in gene expression. In African trypanosomes, this post-transcriptional modification is detected in hundreds of transcripts and it affects the stability of the variant surface glycoprotein (VSG) transcript in the proliferating blood stream form. However, how m6A landscape varies across the life cycle remains poorly defined. Using full-length, non-fragmented RNA, we immunoprecipitated and sequenced m6A-modified transcripts across three life cycle stages ofTrypanosoma brucei– slender (proliferative), stumpy (quiescent), and procyclic forms (proliferative). We found that 1037 transcripts are methylated in at least one of these three life cycle stages. While 21% of methylated transcripts are common in the three stages of the life cycle, globally each stage has a distinct methylome. Interestingly, 47% of methylated transcripts are detected in the quiescent stumpy form only, suggesting a critical role for m6A when parasites exit the cell cycle and prepare for transmission by the Tsetse fly. In this stage, we found that a significant proportion of methylated transcripts encodes for proteins involved in RNA metabolism, which is consistent with their reduced transcription and translation. Moreover, we found that not all major surface proteins are regulated by m6A, as procyclins are not methylated, and that, within the VSG repertoire, not all VSG transcripts are demethylated upon parasite differentiation to procyclic form. This study reveals that the m6A regulatory landscape is specific to each life cycle stage, becoming more pervasive asT. bruceiexits the cell cycle.SummaryAfrican trypanosome parasites adapt to mammalian and insect hosts by adjusting gene expression, morphology, and metabolism. In this study, we focus on how N6-methyladenosine (m6A), a post-transcriptional modification, affects the parasite’s transcriptome throughout its differentiation from the mammalian host to the fly. We found that methylation is differentially regulated as the life cycle progresses, being particularly prevalent in the non-proliferative stumpy form, as more methylated transcripts are found at this insect-infective stage than in slender and procyclic forms. We further show that the not all parasite surface proteins are regulated by m6A and that the previously identified link between m6A methylation and the expression level of the major surface protein of bloodstream forms applies to the active variant surface glycoprotein, but not always to silent genes, suggesting two distinct regulatory mechanisms of (de)methylation.

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