L'abordatge de l'assetjament escolar des de la mediació en la Justícia Juvenil de Catalunya

<div><p>The number of paralogs of proteins involved in translation initiation is larger in trypanosomes than in yeasts or many metazoan and includes two poly(A) binding proteins, PABP1 and PABP2, and four eIF4E variants. In many cases, the paralogs are individually essential and are thus unlikely to have redundant functions although, as yet, distinct functions of different isoforms have not been determined. Here, trypanosome PABP1 and PABP2 have been further characterised. PABP1 and PABP2 diverged subsequent to the differentiation of the Kinetoplastae lineage, supporting the existence of specific aspects of translation initiation regulation. PABP1 and PABP2 exhibit major differences in intracellular localization and distribution on polysome fractionation under various conditions that interfere with mRNA metabolism. Most striking are differences in localization to the four known types of inducible RNP granules. Moreover, only PABP2 but not PABP1 can accumulate in the nucleus. Taken together, these observations indicate that PABP1 and PABP2 likely associate with distinct populations of mRNAs. The differences in localization to inducible RNP granules also apply to paralogs of components of the eIF4F complex: eIF4E1 showed similar localization pattern to PABP2, whereas the localisation of eIF4E4 and eIF4G3 resembled that of PABP1. The grouping of translation initiation as either colocalizing with PABP1 or with PABP2 can be used to complement interaction studies to further define the translation initiation complexes in kinetoplastids.</p> </div

Duplication and divergence of PABPs has occurred on multiple occasions during eukaryote evolution. A)

<p>Phylogenetic reconstruction of PABP evolutionary history using predicted protein sequences from four eukaryotic supergroups <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054004#pone.0054004-Adl1" target="_blank">[78]</a>. Phylogenetic analysis was performed with MrBayes, RAxML and PhyML locally, using the WAG model for ML and mixed parameters for MrBayes <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054004#pone.0054004-Ronquist1" target="_blank">[79]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054004#pone.0054004-Guindon1" target="_blank">[80]</a>, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054004#pone.0054004-Stamatakis1" target="_blank">[81]</a>. The tree was generated using the MrBayes topology. Numbers indicate Baysian posterior probabilities and bootstrap support for PhyML and RAxML respectively; a dash indicates a feature not reconstructed under a specific algorithm. Leaves are colorized according to supergroup: green for Viridiplantae, light blue for Opistokhonta, orange for Chromalveolates and purple for Excavata. Taxa are indicated by a two letter code based on Linnean nomenclature; Hs; <i>Homo sapiens</i>, Dm; <i>Drosophila melanogaster</i>, Ce; <i>Caenorhabditis elegans</i>, DR; <i>Danio rerio</i>, Nv; <i>Nematostella vectensis</i>, Cn; <i>Cryptococcus neoformans</i>, Sp; <i>Schizosaccharomyces pombe</i>, An; <i>Aspergillus nidulans</i>, Ro; <i>Rhizopus oryzae</i>, At; <i>Arabidopsis thaliana</i>, Ot; <i>Oryza sativa</i>, Pt; <i>Populus trichocarpa</i>, Pp; <i>Physcomitrella patens</i>, Ot; <i>Ostreococcus tauri</i>, Pf; <i>Plasmodium falciparum</i>, Tg; <i>Toxoplasma gondii</i>, Cp; <i>Cryptosporidium parvum</i>, Tp; <i>Theileria parva</i>, Pr; <i>Phytophthora ramorum</i>, Tb; <i>Trypanosoma brucei</i> and Lm; <i>Leishmania major</i>. Note that he apparent split of the Viridiplantae clade is an artefact of the manner in which the tree has been drawn, and that monophyly is well supported. <b>B)</b> Phylogenetic reconstruction of PABPs within the Excavata prepared as above. Taxa are indicated by a three letter code based on Linnean nomenclature: Tva, <i>Trichomonas vaginalis</i>; Egr, <i>Euglena gracilis</i>; Tbo, <i>Trypanoplasma borrelli</i>; Bsa, <i>Bodo saltans</i>; Pse, <i>Phytomonas serpens</i>; Pda, <i>Phytomomas davidii</i>; Lb, <i>Leishmania braziliensis</i>; Lin, <i>Leishmania infantum</i>; Lma, <i>Leishmania major</i>; Tbr, <i>Trypanosoma brucei</i>; Tco, <i>Trypanosoma congolense</i>; Tcr, <i>Trypanosoma cruzi</i>; Tth, <i>Trypanosoma theileri</i>; Tgr, <i>Trypanosoma grayi</i> and Tca, <i>Trypanosoma carassii</i>.</p

Differential localization of translation initiation factors to inducible RNA granules.

<p><b>A)</b> eYFP fusions of eIF4E1-4 with mChFP-DHH1. <b>B)</b> eIF4G3-eYFP and mChFP-DHH1. <b>C)</b> PABP2-eYFP and eIF4E1-mChFP.</p

Differential localization of PABP1 and PABP2 to inducible RNP granules.

<p>Z-stack projections of fluorescent microscopy images of cells coexpressing PABP1-eYFP and PAPB2-mChFP, PABP1-eYFP and mChFP-DHH1 or PABP2-eYFP and mChFP-DHH1 are shown. Cells were either untreated or treated with nutrient starvation (2 h PBS), heat shock or sinefungin. The arrows point to heat shock induced PABP1 and PABP2 granules that colocalize. Note that for the heat shock experiment of the cell line expressing PABP1 and DHH1 fusions a cell line was used expressing PABP1-mChFP and eYFP-DHH1; images are shown in false colors for clarity.</p

The predicted NLS of PABP2 is not an NLS. A)

<p>Fluorescent microscopy images of cells expressing PABP1-eYFP/PABP2-mChFP either untreated or treated with sinefungin (2 µg/ml), heat shock (41°C) or both sinefungin and heat shock for two hours. <b>B)</b> Alignment of TbPABP1, TbPABP2 and ScPAB1p (region between RRM 3 and 4). The predicted NLS of TbPABP2 is highlighted, as well as the NLS of the yeast protein PAB1p <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0054004#pone.0054004-Brune1" target="_blank">[74]</a>. <b>C and D)</b> The predicted NLS of <i>T. brucei</i> PABP2 (RLRRER) was exchanged with the equivalent region from PABP1 (ALRQKY) (<b>C</b>) and vice versa (<b>D</b>). Both mutant proteins had cytoplasmic localization identical to the wild type proteins (not shown). Fluorescent microscopy images of untreated cells as well as cells treated with sinefungin, heat shock or both are shown.</p

Summary of localization to granules and co-localization shown in this study.

1<p>*low expression levels.</p>2<p>*localization to PBS granules at over-expression.</p

Distribution of TbPABP1 and TbPABP2 across sucrose gradients.

<p>Lysates prepared from a cell line expressing PABP1-4Ty1 treated as indicated were fractionated by sucrose gradient centrifugation. PABP1-4Ty1 and PABP2, as well as the control proteins BiP and P0 were detected across the fractions of the gradients by western blotting. The absorption profile of the sucrose gradient at 254 nm, the western blots and western blot quantifications are shown.</p

Změna základního tónu řeči a transformace hlasu pomocí PSOLA

In the paper a voice transformation approach based on the application of the Pitch Synchronous OverLap and Add /PSOLA/ principle and resampling is proposed. This algorithm has lower computational demands than frequency domain methods