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Differential gene expression in ap2-g and ap2-g2 mutants of Plasmodium berghei

ABSTRACT: Commitment to and completion of sexual development are essential for malaria parasites to be transmitted through mosquitoes. The molecular mechanism(s) responsible for these processes however, remain largely unknown. We have identified two transcription factors (both belonging to the AP2 family) essential for gametocytogenesis. AP2-G mutants are characterised by a complete inability to produce gametocytes. In AP2-G2 mutants the gametocytaemia is very significantly reduced but not completely abolished. We have performed the microarray experiments in order to cokmpare the transcriptomes of these mutantnts to the WT parasites and between each other. As P.berghei parasites are characterised by asynchronous development in the rodent host, the different stage composition of the sample would impact the analysis. Therefore parasites were harvested and matured in in vitro to the schizont stage Pairwaise comparison between the mutants and parental line was performed. 3 biological replicates of each condition were used.

ORGANISM(S): Plasmodium berghei

SUBMITTER: Oliver Billker

PROVIDER: E-GEOD-52859 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Differential gene expression in ap2-g and ap2-g2 mutants of Plasmodium berghei

Project description:Commitment to and completion of sexual development are essential for malaria parasites to be transmitted through mosquitoes. The molecular mechanism(s) responsible for these processes however, remain largely unknown. We have identified two transcription factors (both belonging to the AP2 family) essential for gametocytogenesis. AP2-G mutants are characterised by a complete inability to produce gametocytes. In AP2-G2 mutants the gametocytaemia is very significantly reduced but not completely abolished. We have performed the microarray experiments in order to cokmpare the transcriptomes of these mutantnts to the WT parasites and between each other. As P.berghei parasites are characterised by asynchronous development in the rodent host, the different stage composition of the sample would impact the analysis. Therefore parasites were harvested and matured in in vitro to the schizont stage

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Plasmodium transcription repressor AP2-O3 regulates sex-specific identity of gene expression in female gametocytes [RNA-seq]

Project description:Male and female gametocytes are sexual precursor cells essential for transmission of malaria parasite in the mosquitoes. Differentiation of gametocytes to fertile gametes (gametogenesis) relies on the gender-specific transcriptome. However, how the parasites establish distinct repertoire of gene transcription in the male and female gametocytes remains largely unknown. Here, we report that an Apetala2 (AP2) family transcription factor (TF) AP2-O3 operates as a transcription repressor regulating female gametocyte transcriptome. AP2-O3 is specifically localized in the nucleus of the female gametocytes. AP2-O3-deficient parasites produce apparently normal female gametocytes, which fail to differentiate to fully fertile female gametes, leading to developmental arrest in fertilization and early development post-fertilization. AP2-O3 disruption causes massive up-regulation of transcriptionally dormant male genes and simultaneously down-regulation of highly transcribed female genes in female gametocytes. ChIP-seq and EMSA analysis establish AP2-O3 as a transcription repressor that targets a significant proportion of the upregulated male genes by recognizing an eight-base DNA motif in the promoters. In addition, the maternal AP2-O3 is removed after fertilization, which is required for the zygote to ookinete development. These results demonstrate that global transcriptional repression of male genes in the female gametocytes is required for safeguarding female-specific transcriptome and essential for the mosquito transmission of Plasmodium.

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Project description:AP2-G is a transcription factor (TF) that is essential for gametocytogenesis of malaria parasites; however, the mechanisms for its inducing the cell linage for sexual reproduction remain unclear. In this paper, this problem is addressed by investigating its expression profile and by determining target genes genome-wide in Plasmodium berghei.

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Project description:During the malaria infection, Plasmodium parasites invade the host’s red blood cells where they can differentiate into two different life forms. The majority will replicate asexually and infect new erythrocytes. A small percentage, however, will transform into gametocytes – a specialized sexual stage able to survive and develop when taken up by Anopheles mosquito. As the gametocytes ensure the parasite’s transmission to a new host, their generation is an attractive target for new antimalarial interventions. The molecular mechanisms controlling gametocytogenesis, however, remain largely unknown due to the technical challenges: the early gametocytes are morphologically indistinguishable from asexual parasites and present in very low numbers during the infection. Recently, AP2-G - a transcription factor from an apicomplexa-specific apiAP2 family – was described as indispensable for gametocyte commitment in both human malaria parasite Plasmodium falciparum and rodent malaria model Plasmodium berghei. Therefore, we have decided to test whether the overexpression of this factor alone could increase gametocyte production and enable the investigation of uncharacterised, earliest stages of gametocyte development. To this end, we have engineered PBGAMi - a Plasmodium berghei line, in which all parasites were ap2-g deficient by default but able to overexpress it when induced with rapamycin. While the control parasites (PBGAMi R-), as expected, differentiated into asexual forms (schizonts) only, almost all rapamycin-treated parasites (PBGAMi R+) transformed into gametocytes. We used the generated line to perform RNA-seq analysis of the R- and R+ populations at different time points of their development and identify the changes arising between them, mapping the sequence of events leading to the formation of gametocytes.

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