Antiprotozoische Struktur‐Aktivitäts‐Beziehungen von synthetischen Leucinostatin‐Derivaten und Aufklärung ihres Wirkprinzips

Leucinostatin A ist eine der potentesten antiprotozoischen Verbindungen, die jemals beschrieben wurden, aber bisher war wenig über die Struktur-Aktivitäts-Beziehung (SAR) bekannt. Trypanosoma brucei wurde als Protozoen-Modellorganismus verwendet, um synthetisch modifizierte Derivate zu testen. Dabei wurden die vereinfachten, aber gleichermaßen aktiven Verbindungen 2 (ZHAWOC6025) und 4 (ZHAWOC6027) identifiziert. Anschließend wurden Modifikationen in allen Teilen des Moleküls durchgeführt, um ein besseres SAR-Verständnis zu erlangen. Die antiprotozoische SAR stimmte mit der SAR in Phospholipid-Liposomen überein, wobei die Membranintegrität, das Durchlässigkeitsverhalten und die Dynamik untersucht wurden. Die antiprotozoale Wirkung der natürlichen und synthetischen Leucinostatine liegt in der Destabilisierung der inneren Mitochondrienmembran, wie durch Ultrastrukturanalyse, Elektronenmikroskopie und Mitochondrienfärbung gezeigt wurde. Eine subletale Langzeitexposition von T. brucei (200 Passagen) und ein siRNA-Screening von 12′000 Mutanten zeigten keine Anzeichen einer Resistenzentwicklung gegenüber den synthetischen Derivaten

Oligosaccharyltransferase Inhibition Induces Senescence in RTK-Driven Tumor Cells

Asparagine (N)-linked glycosylation is a protein modification critical for glycoprotein folding, stability, and cellular localization. To identify small molecules that inhibit new targets in this biosynthetic pathway, we initiated a cell-based high throughput screen and lead compound optimization campaign that delivered a cell permeable inhibitor (NGI-1). NGI-1 targets the oligosaccharyltransferase (OST), a hetero-oligomeric enzyme that exists in multiple isoforms and transfers oligosaccharides to recipient proteins. In non-small cell lung cancer cells NGI-1 blocks cell surface localization and signaling of the EGFR glycoprotein, but selectively arrests proliferation in only those cell lines that are dependent on EGFR (or FGFR) for survival. In these cell lines OST inhibition causes cell cycle arrest accompanied by induction of p21, autofluorescence, and changes in cell morphology; all hallmarks of senescence. These results identify OST inhibition as a potential therapeutic approach for treating receptor tyrosine kinase-dependent tumors and provides a chemical probe for reversibly regulating N-linked glycosylation in mammalian cells

Skillnader i inomartsvariation i morfologiska karaktärer av Empetrum hermaphroditum mellan habitat

Data i denna studie är baserad på en tidigare studie som undersökt om snödjup påverkar medelvärdet för tillväxt och reproduktion hos Empetrum hermaphroditum över en latitud- och klimatgradient (Bienau et al. 2014). Jag testade effekterna för region och snödjup på intraspecifik variation av tillväxtrelaterade variabler istället, för att klargöra om arten har möjlighet att klara av förändringar i snödjup och ökad växtsäsong i framtiden. Tidigare forskning menar att inomartsvariation beror på resurser i habitatet och väcker frågeställningen om det är större variation hos Empetrum i gynnsamma habitat som björkskog och habitat med tjockt snötäcke än i ogynnsamma habitat som de med tunt snötäcke. Vid analys av tidigare samlad data fanns signifikanta skillnader i variation mellan habitat i några morfologiska karaktärer. Däremot är det inte alla dessa karaktärer som följer det förväntade mönstret om att de gynnsamma habitaten skulle ha större variation. Sammantaget visade analysen signifikant skillnad rörande huvudskotten och sidoskottens längd, levande blad på huvudskotten och torrvikten på stammen. Dessa resultat medför att hypotesen stämmer för några tillväxtrelaterade variabler. This study is based on data of a previous study investigating whether snow depth affects average growth and reproduction of Empetrum hermaphroditum over a latitudinal and climatic gradient (Bienau et al. 2014). I tested the effects region and snow depth on intraspecific variation of growth-related variables instead, to clarify whether the species has the potential to cope with changes in snow depth and increased growing season in the future. Earlier research results led to the hypothesis that intraspecific variation depends on resources in the habitat and raises the question of whether there is higher variation in Empetrum in favorable habitats such as birch forests and habitats with deep snow cover than in adverse habitats such as those with a shallow snow cover. My analyses suggest that there were significant differences in variability between habitats in some morphological characters. However, not all of these characters follow the expected pattern that the favorable habitats would have a greater variety. Overall, significant differences were found in variation in the length of the main and the lateral shoots, leaf vitality on the main shoots and the dry weight of the stem. These results imply that the above hypothesis is correct for some growth-related variables.

Phosphatidylethanolamine is the precursor of the ethanolamine phosphoglycerol moiety bound to eukaryotic elongation factor 1A

In addition to its conventional role during protein synthesis, eukaryotic elongation factor 1A is involved in other cellular processes. Several regions of interaction between eukaryotic elongation factor 1A and the translational apparatus or the cytoskeleton have been identified, yet the roles of the different post-translational modifications of eukaryotic elongation factor 1A are completely unknown. One amino acid modification, which so far has only been found in eukaryotic elongation factor 1A, consists of ethanolamine-phosphoglycerol attached to two glutamate residues that are conserved between mammals and plants. We now report that ethanolamine-phosphoglycerol is also present in eukaryotic elongation factor 1A of the protozoan parasite Trypanosoma brucei, indicating that this unique protein modification is of ancient origin. In addition, using RNA-mediated gene silencing against enzymes of the Kennedy pathway, we demonstrate that phosphatidylethanolamine is a direct precursor of the ethanolamine-phosphoglycerol moiety. Down-regulation of the expression of ethanolamine kinase and ethanolamine-phosphate cytidylyltransferase results in inhibition of phosphatidylethanolamine synthesis in T. brucei procyclic forms and, concomitantly, in a block in glycosylphosphatidylinositol attachment to procyclins and ethanolamine-phosphoglycerol modification of eukaryotic elongation factor 1A

TbLpn, a key enzyme in lipid droplet formation and phospholipid metabolism, is essential for mitochondrial integrity and growth of Trypanosoma brucei.

Mammalian phosphatidic acid phosphatases, also called lipins, show high amino acid sequence identity to Saccharomyces cerevisiae Pah1p and catalyze the dephosphorylation of phosphatidic acid (PA) to diacylglycerol. Both the substrate and product of the reaction are key precursors for the synthesis of phospholipids and triacylglycerol (TAG). We now show that expression of the Trypanosoma brucei lipin homolog TbLpn is essential for parasite survival in culture. Inducible down-regulation of TbLpn in T. brucei procyclic forms increased cellular PA content, decreased the numbers of lipid droplets, reduced TAG steady-state levels and inhibited in vivo [ H]TAG formation after labeling trypanosomes with [ H]glycerol. In addition, fluorescence and transmission electron microscopy revealed that depletion of TbLpn caused major alterations in mitochondrial morphology and function, i.e., the appearance of distorted mitochondrial matrix, and reduced ATP production via oxidative phosphorylation. Effects of lipin depletion on mitochondrial integrity have previously not been reported. N- and C-terminally tagged forms of TbLpn were localized in the cytosol