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search.filters.applied.f.access: openAccess × End date: 2019 × Start date: 2019 × End date: 2015 × Start date: 2012 × Type: Text × Publisher: London : Nature Publishing Group × WGL Institute: ISAS ×

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    Raf kinases mediate the phosphorylation of eukaryotic translation elongation factor 1A and regulate its stability in eukaryotic cells
    (London : Nature Publishing Group, 2012) Sanges, C.; Scheuermann, C.; Zahedi, R.P.; Sickmann, A.; Lamberti, A.; Migliaccio, N.; Baljuls, A.; Marra, M.; Zappavigna, S.; Reinders, J.; Rapp, U.; Abbruzzese, A.; Caraglia, M.; Arcari, P.
    We identified eukaryotic translation elongation factor 1A (eEF1A) Raf-mediated phosphorylation sites and defined their role in the regulation of eEF1A half-life and of apoptosis of human cancer cells. Mass spectrometry identified in vitro S21 and T88 as phosphorylation sites mediated by B-Raf but not C-Raf on eEF1A1 whereas S21 was phosphorylated on eEF1A2 by both B- and C-Raf. Interestingly, S21 belongs to the first eEF1A GTP/GDP-binding consensus sequence. Phosphorylation of S21 was strongly enhanced when both eEF1A isoforms were preincubated prior the assay with C-Raf, suggesting that the eEF1A isoforms can heterodimerize thus increasing the accessibility of S21 to the phosphate. Overexpression of eEF1A1 in COS 7 cells confirmed the phosphorylation of T88 also in vivo. Compared with wt, in COS 7 cells overexpressed phosphodeficient (A) and phospho-mimicking (D) mutants of eEF1A1 (S21A/D and T88A/D) and of eEF1A2 (S21A/D), resulted less stable and more rapidly proteasome degraded. Transfection of S21 A/D eEF1A mutants in H1355 cells increased apoptosis in comparison with the wt isoforms. It indicates that the blockage of S21 interferes with or even supports C-Raf induced apoptosis rather than cell survival. Raf-mediated regulation of this site could be a crucial mechanism involved in the functional switching of eEF1A between its role in protein biosynthesis and its participation in other cellular processes.
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    The bruchpilot cytomatrix determines the size of the readily releasable pool of synaptic vesicles
    (London : Nature Publishing Group, 2013) Matkovic, Tanja; Siebert, Matthias; Knoche, Elena; Depner, Harald; Mertel, Sara; Owald, David; Schmidt, Manuela; Thomas, Ulrich; Sickmann, Albert; Kamin, Dirk; Hell, Stefan W.; Bürger, Jörg; Hollmann, Christina; Mielke, Thorsten; Wichmann, Carolin; Sigrist, Stephan J.
    Synaptic vesicles (SVs) fuse at a specialized membrane domain called the active zone (AZ), covered by a conserved cytomatrix. How exactly cytomatrix components intersect with SV release remains insufficiently understood. We showed previously that loss of the Drosophila melanogaster ELKS family protein Bruchpilot (BRP) eliminates the cytomatrix (T bar) and declusters Ca2+ channels. In this paper, we explored additional functions of the cytomatrix, starting with the biochemical identification of two BRP isoforms. Both isoforms alternated in a circular array and were important for proper T-bar formation. Basal transmission was decreased in isoform-specific mutants, which we attributed to a reduction in the size of the readily releasable pool (RRP) of SVs. We also found a corresponding reduction in the number of SVs docked close to the remaining cytomatrix. We propose that the macromolecular architecture created by the alternating pattern of the BRP isoforms determines the number of Ca2+ channel-coupled SV release slots available per AZ and thereby sets the size of the RRP.
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    Biological pathways modulated by antipsychotics in the blood plasma of schizophrenia patients and their association to a clinical response
    (London : Nature Publishing Group, 2015) Martins-de-Souza, Daniel; Solari, Fiorella A.; Guest, Paul C.; Zahedi, René P.; Steiner, Johann
    Proteomics is a valuable tool to unravel molecular mechanisms involved in human disorders. Considering the mediocre effectiveness of antipsychotics, which are the main class of drug used to treat schizophrenia, we analyzed a cohort of 58 schizophrenia patients who had blood collected before and after 6 weeks of antipsychotic treatment using a shotgun mass spectrometry proteomic profiling approach. Our aim was to unravel molecular pathways involved with an effective drug response. The results showed that all patients had essentially the same biochemical pathways triggered Independent of the antipsychotic response outcome. However, we observed that these pathways were regulated in different directions in blood samples from those who responded well to antipsychotics, compared with those who had a poorer outcome. These data are novel, timely and may help to guide new research efforts in the design of new treatments or medications for schizophrenia based on biologically relevant pathways.