The <i>T. cruzi</i> mRNA population is partially accumulated into the nucleolus in response to severe heat shock.

<p>Localization of the mRNA population using either an (A) oligo(dT) or (B) mini-exon probe in untreated, heat-shocked and recovered <i>T. cruzi</i> epimastigotes. In addition, cells were pre-treated with RNase A before performing FISH (bottom panels). Nuclei were counterstained with DAPI (blue). The white arrows indicate the nucleolar localization for both probes. (C) RNA FISH using a Cy3-labelled oligo(dA)30 probe in untreated parasites and parasites exposed to 40°C for 2 h. N: nucleus, K: kinetoplast, Nu: nucleolus. Size bars represent 2 µm. Representative nuclei are shown. (D) A quantitative analysis of experiments is shown in panels (A) and (B). The results are expressed as mean +/− SD from at least three independent experiments (a minimum of 100 parasites per experiment were counted).</p

Poly(A)+ RNA in response to heat shock is located in cytoplasmic granules but not into the nucleolus in <i>T. brucei</i>.

<p>Poly(A)+ RNA subcellular localization in <i>T. brucei</i> procyclic forms (A) untreated, (B) exposed to heat shock at 40°C for 2 h, (C) pre-treated with RNase A after heat shock, and (D) incubated with ActD for 4 h. The white arrows indicate the nucleolus. Nuclei were counterstained with DAPI (blue). N: nucleus, K: kinetoplast, Nu: nucleolus. Size bars represent 2 µm. Representative parasites are shown.</p

Hsp70 mRNA is able to bypass the nucleolar accumulation.

<p>FISH analysis for (A) α-Tub, (B) Smug and (C) Hsp70 mRNAs in parasites subjected to heat shock at 40°C for 2 h. In addition, cells were pre-treated with RNase A before performing FISH (bottom panels). The white arrows indicate the nucleolus. Nuclei were counterstained with DAPI (blue). Size bars represent 2 µm. Representative nuclei are shown. (D) A quantitative analysis of the experiments shown in panels (A), (B) and (C). The results are expressed as mean +/− SD from at least three independent experiments (a minimum of 100 parasites per experiment were counted).</p

Integrative model showing the behaviour of RBPs and mRNAs in response to severe heat shock in <i>T. cruzi</i>.

<p>Under normal conditions, the mRNAs are exported to the cytoplasm to be target either to translation, storage or degradation. When epimastigotes are exposed to severe heat shock, there is a decrease in transcription which likely triggers the nucleolar accumulation of certain RBPs (for instance, TcSR62 and TcPTB2). In addition, mRNAs are relocalized into the nucleolus where they might be stored/protected until favourable environmental conditions are resumed. On the other hand, the Hsp70 mRNA could bypass such nucleolar retention, being transported to the cytoplasm where its translation can continue even under severe heat shock. Unidentified, but probable, factors/events are indicated with a question mark. See the <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0043715#s3" target="_blank">discussion</a> section for details. SL, spliced leader.</p