Serial Analysis of Gene Expression in Plasmodium falciparum Reveals the Global Expression Profile of Erythrocytic Stages and the Presence of Anti-Sense Transcripts in the Malarial Parasite

Abstract

Serial analysis of gene expression (SAGE) was applied to the malarial parasite Plasmodium falciparum to characterize the comprehensive transcriptional profile of erythrocytic stages. A SAGE library of ϳ8335 tags representing 4866 different genes was generated from 3D7 strain parasites. Basic local alignment search tool analysis of high abundance SAGE tags revealed that a majority (88%) corresponded to 3D7 sequence, and despite the low complexity of the genome, 70% of these highly abundant tags matched unique loci. Characterization of these suggested the major metabolic pathways that are used by the organism under normal culture conditions. Furthermore several tags expressed at high abundance (30% of tags matching to unique loci of the 3D7 genome) were derived from previously uncharacterized open reading frames, demonstrating the use of SAGE in genome annotation. The open platform "profiling" nature of SAGE also lead to the important discovery of a novel transcriptional phenomenon in the malarial pathogen: a significant number of highly abundant tags that were derived from annotated genes (17%) corresponded to antisense transcripts. These SAGE data were validated by two independent means, strand specific reverse transcription-polymerase chain reaction and Northern analysis, where antisense messages were detected in both asexual and sexual stages. This finding has implications for transcriptional regulation of Plasmodium gene expression.

Chromatin associated sense and antisense noncoding RNAs are transcribed from the var gene family of virulence genes of the malaria parasite Plasmodium falciparum

“…In much the same way, sterile transcripts may direct chromatin-associated factors to target specific var genes for silencing. Given the data presented here, the recent identification in silico of more than 30 novel ncRNAs of unknown function (Chakrabarti et al 2007;Mourier et al 2008) and evidence that 17% of annotated var noncoding RNAs www.rnajournal.org 123 genes express detectable antisense RNA transcripts, it is likely that RNA-mediated gene regulation is an integral part of P. falciparum biology (Patankar et al 2001).…”

Monitoring the chromosome 2 intraerythrocytic transcriptome of Plasmodium falciparum using oligonucleotide arrays.

“…We also compared our results with those obtained with SAGE analysis (D. Wirth personal communication, Harvard), and results were generally in agreement (data not shown). 10 To verify these array results using conventional methods, we performed semiquantitative RT-PCR, as well as real time RT-PCR and Northern blot analysis on a subset of genes that were expressed at low levels, moderate levels, or high levels in the schizont stage of the life cycle. (Figure 3).…”

Chloroquine Resistance Modulated in Vitro by Expression Levels of the Plasmodium falciparum Chloroquine Resistance Transporter

“…However, concerns remain because of the lack of critical RNA interference proteins in the P. falciparum genome (64), including the RNAdependent RNA polymerase Dicer. Alternative forms of RNAdependent gene regulation may nonetheless exist in P. falciparum, as evidenced by the discovery of unique RNA-protein interactions that were found to affect antimalarial antifolate susceptibility (65) as well as the finding of abundant antisense transcripts in this organism (66). An improved understanding of P. falciparum gene regulation and its application to the study of drug resistance and pathogenesis can facilitate efforts to translate post-genomic research into practical solutions to combat malaria.…”