Real-time in vivo Global Transcriptional Dynamics During Plasmodium falciparum Blood-stage Development

Abstract

ABSTRACT:Genome-wide analysis of transcription in the human malaria parasite Plasmodium falciparum has revealed robust variation in steady-state mRNA abundance throughout the 48-hour intraerythrocytic developmental cycle (IDC) suggesting that this process is highly dynamic and tightly regulated. However, the precise timing of mRNA transcription and decay remains poorly understood due to the utilization of methods that only measure total RNA and cannot differentiate between newly transcribed, decaying and stable cellular RNAs. Here we utilize rapid 4-thiouracil (4-TU) incorporation via pyrimidine salvage to specifically label, capture and quantify newlysynthesized P. falciparum RNA transcripts at every hour throughout the IDC following erythrocyte invasion. This high resolution global analysis of the transcriptome captures the timing and rate of transcription for each newly synthesized mRNA in vivo, revealing active transcription throughout all stages of the IDC. To determine the fraction of active transcription and/or transcript stabilization contributing to the total mRNA abundance at each timepoint we have generated a statistical model to fit the data for each gene which reveals varying degrees of transcription and stabilization for each mRNA corresponding to developmental transitions and independent of abundance profile. Finally, our results provide new insight into co-regulation of mRNAs throughout the IDC through regulatory DNA sequence motifs associated with these processes, thereby expanding our understanding of P. falciparum mRNA dynamics. INTRODUCTION:

Refining the transcriptome of the human malaria parasite Plasmodium falciparum using amplification-free RNA-seq

“…The P. falciparum genome is 23.3 Mb in size and encodes over 5400 genes [3]. Most parasite genes are transcriptionally regulated during the IDC, often expressed across multiple time points but with a single peak of maximum abundance per gene [4][5][6]. Another study has compared the IDC transcriptome profiles of three laboratory strains (3D7, HB3 and Dd2; with origins in West Africa, Latin America and Asia, respectively), demonstrating that gene expression was remarkably conserved between strains from across the globe, despite the strains being isolated at different times from disparate geographical locations [7].…”

Global Repertoire of Human Antibodies Against Plasmodium falciparum RIFINs, SURFINs, and STEVORs in a Malaria Exposed Population

“…In the reference P. falciparum 3D7 genome, RIFIN is the largest multigene family and is encoded by ∼180 rif genes (7). Multiple RIFINs are simultaneously expressed 11-12 h post RBC invasion (8) and are subsequently exported to the surface of iRBCs. Some RIFIN variants are expressed in other parasite stages such as merozoites and sporozoites (9,10).…”

Hierarchical transcriptional control regulates Plasmodium falciparum sexual differentiation

“…Therefore, gametocytogenesis relies on a more specialized program of gene expression compared to asexual development, with only 45% of transcripts showing increased abundance during gametocyte development ( Fig. 2a) compared to the 80-95% of transcripts increased during specific phases of asexual development [11,19,45]. Interestingly, individual clusters showed specific patterns of gene expression throughout gametocyte development ( Fig.…”

“…These patterns have been investigated to some extent for asexual replication where P. falciparum parasites use both transcriptional [6][7][8] and post-transcriptional processes [9,10] to effect a cascade of coordinated, stage-specific gene expression [11,12]. Despite the identification of some putative regulators of gene expression, including the Apicomplexan-specific ApiAP2 family of transcription factors [13][14][15] and epigenetic regulation of particular gene families [16,17], the specific mechanisms controlling transcriptional activation in the parasite are incompletely understood, with recent data clearly showing mRNA dynamics are also influenced by additional post-transcriptional mechanisms [18,19].…”