Stochastic versus stable transcriptional differences on Plasmodium falciparum DNA microarrays

Ganesan, Karthikeyan; Jiang, Lei; Rathod, Pradipsinh K.

doi:10.1016/s0020-7519(02)00182-0

Cited by 9 publications

(10 citation statements)

References 48 publications

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“…The biological significance of this observation is unclear, but inconsistent MPSS signatures present in only a single run have been observed from genes in other organisms (32). Such differences may be stochastic rather than stable and the expression profile could differ at another time point, or in different cell lines, as observed previously with P.falciparum (33). Additional data relating to expression of T.parva telomere-associated ORFs provide support for this hypothesis.…”

Section: Discussionmentioning

confidence: 78%

Analysis of the transcriptome of the protozoan Theileria parva using MPSS reveals that the majority of genes are transcriptionally active in the schizont stage

Bishop

Shah

Pellé

et al. 2005

Nucleic Acids Research

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Massively parallel signature sequencing (MPSS) was used to analyze the transcriptome of the intracellular protozoan Theileria parva. In total 1 095 000, 20 bp sequences representing 4371 different signatures were generated from T.parva schizonts. Reproducible signatures were identified within 73% of potentially detectable predicted genes and 83% had signatures in at least one MPSS cycle. A predicted leader peptide was detected on 405 expressed genes. The quantitative range of signatures was 4–52 256 transcripts per million (t.p.m.). Rare transcripts (<50 t.p.m.) were detected from 36% of genes. Sequence signatures approximated a lognormal distribution, as in microarray. Transcripts were widely distributed throughout the genome, although only 47% of 138 telomere-associated open reading frames exhibited signatures. Antisense signatures comprised 13.8% of the total, comparable with Plasmodium. Eighty five predicted genes with antisense signatures lacked a sense signature. Antisense transcripts were independently amplified from schizont cDNA and verified by sequencing. The MPSS transcripts per million for seven genes encoding schizont antigens recognized by bovine CD8 T cells varied 1000-fold. There was concordance between transcription and protein expression for heat shock proteins that were very highly expressed according to MPSS and proteomics. The data suggests a low level of baseline transcription from the majority of protein-coding genes.

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Section: Discussionmentioning

confidence: 78%

Analysis of the transcriptome of the protozoan Theileria parva using MPSS reveals that the majority of genes are transcriptionally active in the schizont stage

Bishop

Shah

Pellé

et al. 2005

Nucleic Acids Research

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“…For each time point, samples from biological duplicates were hybridized to four microarray slides, including dye exchanges. The biological duplicates were from independently propagated cultures to minimize misleading, stochastic variations in gene expression for surface proteins ([32], Figure S2 and Figure S3 for description of data normalization and variation between technical and biological variability). In total, the present normalized data is derived from 123 microarray hybridizations.…”

Section: Resultsmentioning

confidence: 99%

“…The present experiments used spotted DNA microarrays [71] that were fabricated as previously described [9],[32]. Commercially available malaria oligonucleotides (Operon, version 1.1, https://www.operon.com/arrays/oligosets_malaria.php) were combined with in-house oligonucleotides representing P. falciparum genes in folate and nucleic acid metabolism.…”

Section: Methodsmentioning

confidence: 99%

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A Genetically Hard-Wired Metabolic Transcriptome in Plasmodium falciparum Fails to Mount Protective Responses to Lethal Antifolates

Ganesan¹,

Ponmee²,

Jiang³

et al. 2008

PLoS Pathog

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Genome sequences of Plasmodium falciparum allow for global analysis of drug responses to antimalarial agents. It was of interest to learn how DNA microarrays may be used to study drug action in malaria parasites. In one large, tightly controlled study involving 123 microarray hybridizations between cDNA from isogenic drug-sensitive and drug-resistant parasites, a lethal antifolate (WR99210) failed to over-produce RNA for the genetically proven principal target, dihydrofolate reductase-thymidylate synthase (DHFR-TS). This transcriptional rigidity carried over to metabolically related RNA encoding folate and pyrimidine biosynthesis, as well as to the rest of the parasite genome. No genes were reproducibly up-regulated by more than 2-fold until 24 h after initial drug exposure, even though clonal viability decreased by 50% within 6 h. We predicted and showed that while the parasites do not mount protective transcriptional responses to antifolates in real time, P. falciparum cells transfected with human DHFR gene, and adapted to long-term WR99210 exposure, adjusted the hard-wired transcriptome itself to thrive in the presence of the drug. A system-wide incapacity for changing RNA levels in response to specific metabolic perturbations may contribute to selective vulnerabilities of Plasmodium falciparum to lethal antimetabolites. In addition, such regulation affects how DNA microarrays are used to understand the mode of action of antimetabolites.

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“…If true, it follows that divergent phenotypes between strains, such as drug resistance, do not result from variation in the transcriptional profile. This interpretation runs contrary to small datasets on direct co-hybridization of cDNA from malaria parasites [3], recent evidence that malaria parasites display distinct physiological states in their in vivo transcriptional profiles [14], and numerous observations that modifications in transcriptional regulation underpin complex adaptations in human, fly, worm, and yeast clonal populations [15–18]. …”

Section: Introductionmentioning

confidence: 85%

Regulatory Hotspots in the Malaria Parasite Genome Dictate Transcriptional Variation

et al. 2008

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The determinants of transcriptional regulation in malaria parasites remain elusive. The presence of a well-characterized gene expression cascade shared by different Plasmodium falciparum strains could imply that transcriptional regulation and its natural variation do not contribute significantly to the evolution of parasite drug resistance. To clarify the role of transcriptional variation as a source of stain-specific diversity in the most deadly malaria species and to find genetic loci that dictate variations in gene expression, we examined genome-wide expression level polymorphisms (ELPs) in a genetic cross between phenotypically distinct parasite clones. Significant variation in gene expression is observed through direct co-hybridizations of RNA from different P. falciparum clones. Nearly 18% of genes were regulated by a significant expression quantitative trait locus. The genetic determinants of most of these ELPs resided in hotspots that are physically distant from their targets. The most prominent regulatory locus, influencing 269 transcripts, coincided with a Chromosome 5 amplification event carrying the drug resistance gene, pfmdr1, and 13 other genes. Drug selection pressure in the Dd2 parental clone lineage led not only to a copy number change in the pfmdr1 gene but also to an increased copy number of putative neighboring regulatory factors that, in turn, broadly influence the transcriptional network. Previously unrecognized transcriptional variation, controlled by polymorphic regulatory genes and possibly master regulators within large copy number variants, contributes to sweeping phenotypic evolution in drug-resistant malaria parasites.

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Stochastic versus stable transcriptional differences on Plasmodium falciparum DNA microarrays

Cited by 9 publications

References 48 publications

Analysis of the transcriptome of the protozoan Theileria parva using MPSS reveals that the majority of genes are transcriptionally active in the schizont stage

Analysis of the transcriptome of the protozoan Theileria parva using MPSS reveals that the majority of genes are transcriptionally active in the schizont stage

A Genetically Hard-Wired Metabolic Transcriptome in Plasmodium falciparum Fails to Mount Protective Responses to Lethal Antifolates

Regulatory Hotspots in the Malaria Parasite Genome Dictate Transcriptional Variation

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