Transcriptional heterogeneity and tightly regulated changes in gene expression during Plasmodium berghei sporozoite development

Abstract

Despite the critical role of Plasmodium sporozoites in malaria transmission, we still know little about the mechanisms underlying their development in mosquitoes. Here, we use single-cell RNA sequencing to characterize the gene expression profiles of 16,038 Plasmodium berghei sporozoites isolated throughout their development from midgut oocysts to salivary glands, and from forced salivation experiments. Our results reveal a succession of tightly regulated changes in gene expression occurring during the maturation of sporozoites and highlight candidate genes that could play important roles in oocyst egress, sporozoite motility, and the mechanisms underlying the invasion of mosquito salivary glands and mammalian hepatocytes. In addition, the single-cell data reveal extensive transcriptional heterogeneity among parasites isolated from the same anatomical site, suggesting that Plasmodium development in mosquitoes is asynchronous and regulated by intrinsic as well as environmental factors. Finally, our analyses show a decrease in transcriptional activity preceding the translational repression observed in mature sporozoites and associated with their quiescent state in salivary glands, followed by a rapid reactivation of the transcriptional machinery immediately upon salivation.

Single-cell RNA sequencing ofPlasmodiumvivax sporozoites reveals stage- and species-specific transcriptomic signatures

“…berghei midgut sporozoites (Bogale et al, 2021;Ruberto et al, 2021). We interpret the detection of these markers in P. vivax to represent sporozoites that have recently invaded the salivary glands.…”

“…Processing, alignment, and pseudo-bulk assessment of P. vivax sporozoite scRNA-seq data Given its high-throughput capability and prior use for sporozoites from murine-infecting Plasmodium species (Bogale et al, 2021;Ruberto et al, 2021), we used the 10X Genomics gene expression platform to profile the transcriptomes of individual P. vivax sporozoites. We dissected and purified sporozoites from the salivary glands of An.…”

“…In total, we found 159 markers (adjusted p-value < 0.05, Table S5). Notably, sporozoite-specific protein, S10 (PVP01_0304200), was a marker for sporozoites in C1 (Figure 3C; Table S5), and has previously been shown to be highly transcribed in midgut sporozoites (Bogale et al, 2021;Ruberto et al, 2021). C2 markers included circumsporozoite protein (PVP01_0835600); early transcribed membrane protein (PVP01_0602100); TRAP-like protein (PVP01_1132600); and sporozoite surface protein essential for liver stage development (PVP01_0938800) (Figure 3C;…”

“…Single-cell RNA sequencing methods (scRNA-seq) constitute a recent advancement applicable for assessing parasite-to-parasite differences. ScRNA-seq has differentiated multiple transcriptomic states among individual P. berghei and P. falciparum sporozoites (Bogale et al, 2021; Howick et al, 2019; Real et al, 2021; Ruberto et al, 2021). However, the extent to which P. vivax sporozoites vary at the single-cell level has not been studied.…”

5-methylcytosine modification byPlasmodiumNSUN2 stabilizes mRNA and mediates the development of gametocytes

“…While this does not rule out the possibility that erasers may yet be found, it is possible that m 5 Control of mRNA turnover and translational repression has been shown to be essential for Plasmodium sexual development and is mediated by a member of the DDX6-family of DEAD box RNA helicases (37). Translational repression also occurs in mature sporozoites and is associated with their quiescence in mosquito salivary glands until blood feeding and salivation (38)(39)(40).…”