<i>Rif</i> transcription and H3K9 modifications after reselection over CHO-CD36 cells.

<p>(A) The <i>rif</i> transcript quantities of freshly CHO-CD36 selected parasites in trophozoite stage (20–24 h post invasion) were monitored by RT-qPCR in relation to the internal control PF070073. (B) Histone modifications were measured by ChIP as described and significant differences in signals between H3K9ac and H3K9me3 modifications are depicted by asterisks (Student's T test, <i>p</i><0.05). ChIP was performed in triplicates.</p

Relative quantities of <i>rif</i> transcripts and chromatin enrichment for H3K9ac, H3K9me3 and H3K4me2 (poised mark) in trophozoite and schizont stages for selected <i>rif</i> loci.

<p>A: Relative transcript amounts of chosen <i>rif</i> targets in trophozoites and schizonts in 3D7 parasites of the same reinvasion, calculated as above. B: Ratios of H3K9ac/me3 enrichment in trophozoites (20–24 h p.i.) and schizonts (30–34 h p.i.). C: Scatter plots of the relative transcription in schizonts plotted against the H3K4me2 enrichment in trophozoites showing a positive and significant correlation (R² = 0.4929, p = 0.02). The ChIP experiment was done in biological triplicates.</p

Transcriptional analysis and chromatin enrichment after 10 and 20 reinvasions.

<p>A: Transcription of 3D7-CD36 parasites of chosen <i>rif</i> targets are shown as relative values (2^−ΔCt) using the gene of plasmodial t-seryl Synthetase (PF070073) as an internal control. B, C: Chromatin enrichment at selected <i>rif</i> loci shown as H3K9ac/H3K9me3 ratios obtained for 20–24 h trophozoite samples after 10 (B) and 20 reinvasions (C). D: Scatter plot showing the relation of histone modifications and transcript quantity. The coefficients of relative transcription at each loci (at 20 and 10 reinvasions) were plotted against the coefficient of H3K9ac/me3 ratios_{20 reinvasions}/H3K9ac/me3 ratios_{10 reinvasions}. The ChIP experiment was done in biological triplicates and Pearson's correlation of the values is indicated (p = 0.03, one tailed probability). Note that the extreme outliers PF080138 and PFB0040c were not considered.</p

Influence of primer localization on the qPCR signal after ChIP at two <i>rif</i> loci.

<p>A: Immunoprecipitated material from the experiment in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029881#pone-0029881-g002" target="_blank">Fig. 2</a> was used and the localization of the employed primer pairs in qPCR is shown for the two tested <i>rif</i> ORFs. B: <i>rif</i> transcription at the two loci was measured as in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029881#pone-0029881-g001" target="_blank">Figure 1</a>. C: The H3K9ac and H3K9me3 signals in ChIP were expressed as ratios. Note that the oligonucleotide primers used for the experiments in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029881#pone-0029881-g001" target="_blank">Figures 1</a>–<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029881#pone-0029881-g002" target="_blank"></a><a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0029881#pone-0029881-g003" target="_blank">3</a> were PFI0025c_3 and PF100397_2. The ChIP experiment was done in biological triplicates.</p