Single Cell RNA Sequencing Reveals Transcriptional Heterogeneity and Stochastic Gene Expression in Plasmodium falciparum Schizonts
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ABSTRACT:
PROVIDER: PRJNA690786 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR13388100_1.fastq.gz | Fastqsanger.gz | |||
| SRR13388100_2.fastq.gz | Fastqsanger.gz | |||
| SRR13388200_1.fastq.gz | Fastqsanger.gz | |||
| SRR13388200_2.fastq.gz | Fastqsanger.gz | |||
| SRR13388300_1.fastq.gz | Fastqsanger.gz |
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Project description:Genetically identical cells are known to exhibit differential phenotypes in the same environmental conditions. These phenotypic variants are linked to transcriptional stochasticity and have been shown to contribute towards adaptive flexibility of a wide range of unicellular organisms such as yeast and bacteria. Here, we investigated transcriptional heterogeneity and stochastic gene expression in P. falciparum by performing single cell RNA sequencing on blood stage schizonts. Our data reveals significant transcriptional variations in the schizonts stage that appear to reflect several diversification processes including putative developmental “checkpoint”; transcriptomically distinct parasite cells sub-populations; transcriptional switches in variable gene families and a distinct group of highly variable transcripts. Most of these features of transcriptional variability were preserved in isogenic parasite cell populations (albeit with a lesser amplitude) suggesting a role of epigenetic factors in cell-to-cell transcriptional variations in human malaria parasites. Finally, to validate our findings, we applied quantitative qRT-PCR and RNA-FISH approach and confirmed stochastic expression of several genes such as msp1, msp3, msp7, rhopH2 and eba181, some of which represent key candidates for invasion blocking vaccines
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Project description:Plasmodium falciparum merozoite surface protein MSPDBL2 is a highly polymorphic target of naturally acquired immune responses encoded by a single copy gene under strong balancing selection in natural populations. The MSPDBL2 protein is expressed in only a minority of mature schizonts of any cultured parasite line, and mspdbl2 gene expression in culture is increased in response to overexpression of the gametocyte development inducer GDV1 in an engineered parasite clone, so there is a need to characterise its natural expression variation. In this study, MSPDBL2 in mature schizonts was analyzed in the first ex vivo culture cycle of 102 clinical isolates from four endemic populations in different West African countries, by immunofluorescence microscopy with antibodies against a conserved region of the protein. In most isolates, less than 1% of mature schizonts were positive for MSPDBL2 (median of 0.6% overall), but the frequency distribution was highly skewed as eleven isolates had more than 3% schizonts positive and one had 73% positive. The expression frequencies were similar across the four endemic populations, and there was no significant difference between the clinical isolates overall and frequencies of positive schizonts previously seen in cultured P. falciparum laboratory lines. To explore whether expression of other gene loci correlated with MSPDBL2 expression, whole transcriptome sequencing was performed on schizont-enriched material from 17 of the clinical isolates with a wide range of proportions of schizonts positive. Transcripts of particular parasite genes were highly significantly positively correlated with MSPDBP2 positivity in schizonts as well as with mspdbl2 gene transcript levels, with overrepresentation of many genes previously implicated as likely to be involved in gametocytogenesis, but not including other genes including the gametocytogenesis master regulator ap2g. Although MSPDBL2 expression occurs in a highly variable proportion of schizonts in clinical isolates, and correlation with expression of some gametocytogenesis-related genes is consistent with regulation by GDV1, it is not apparently a direct marker of sexual commitment and its function in the parasite remains to be determined.
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