Processing of the amyloid precursor protein (APP) by b-and c-secretases leads to the generation of amyloid-b (Ab) peptides with varying lengths. Particularly Ab42 contributes to cytotoxicity and amyloid accumulation in Alzheimer's disease (AD). However, the precise molecular mechanism of Ab42 generation has remained unclear. Here, we show that an amino-acid motif GxxxG within the APP transmembrane sequence (TMS) has regulatory impact on the Ab species produced. In a neuronal cell system, mutations of glycine residues G29 and G33 of the GxxxG motif gradually attenuate the TMS dimerization strength, specifically reduce the formation of Ab42, leave the level of Ab40 unaffected, but increase Ab38 and shorter Ab species. We show that glycine residues G29 and G33 are part of a dimerization site within the TMS, but do not impair oligomerization of the APP ectodomain. We conclude that c-secretase cleavages of APP are intimately linked to the dimerization strength of the substrate TMS. The results demonstrate that dimerization of APP TMS is a risk factor for AD due to facilitating Ab42 production.

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Aims: Staphylococcus aureus is a major human pathogen and has to cope with reactive oxygen and chlorine species (ROS, RCS) during infections, which requires efficient protection mechanisms to avoid destruction. Here, we have investigated the changes in the RNA-seq transcriptome by the strong oxidant sodium hypochlorite (NaOCl) in S. aureus USA300 to identify novel redox-sensing mechanisms that provide protection under infection conditions.Results: NaOCl stress caused an oxidative stress response in S. aureus as indicated by the induction of the PerR, QsrR, HrcA, and SigmaB regulons in the RNA-seq transcriptome. The hypR-merA (USA300HOU_0588-87) operon was most strongly upregulated under NaOCl stress, which encodes for the Rrf2-family regulator HypR and the pyridine nucleotide disulfide reductase MerA. We have characterized HypR as a novel redox-sensitive repressor that controls MerA expression and directly senses and responds to NaOCl and diamide stress via a thiol-based mechanism in S. aureus. Mutational analysis identified Cys33 and the conserved Cys99 as essential for NaOCl sensing, while Cys99 is also important for repressor activity of HypR in vivo. The redox-sensing mechanism of HypR involves Cys33-Cys99 intersubunit disulfide formation by NaOCl stress both in vitro and in vivo. Moreover, the HypR-controlled flavin disulfide reductase MerA was shown to protect S. aureus against NaOCl stress and increased survival in J774A.1 macrophage infection assays.Conclusion and Innovation: Here, we identified a new member of the widespread Rrf2 family as redox sensor of NaOCl stress in S. aureus that uses a thiol/disulfide switch to regulate defense mechanisms against the oxidative burst under infections in S. aureus. Antioxid. Redox Signal. 29, 615–636.

By using degenerate oligonucleotide primers deduced from the conserved regions of the mammalian somatostatin receptors, a novel G-protein-coupled receptor from Drosophila melanogaster has been isolated exhibiting structural similarities to mammalian somatostatin/ galanin/opioid receptors. To identify the bioactive ligand, a 'reverse physiology' strategy was used whereby orphan Drosophila receptor-expressing frog oocytes were screened against potential ligands. Agonistic activity was electrophysiologically recorded as inward potassium currents mediated through co-expressed Gprotein-gated inwardly rectifying potassium channels (GIRK). Using this approach a novel peptide was purified from Drosophila head extracts. Mass spectrometry revealed an octapeptide of 925 Da with a sequence SerArg-Pro-Tyr-Ser-Phe-Gly-Leu-NH 2 reminiscent of insect allatostatin peptides known to control diverse functions such as juvenile hormone synthesis during metamorphosis or visceral muscle contractions. Picomolar concentrations of the synthesized octapeptide activated the cognate receptor response mediated through GIRK1, indicating that we have isolated the 394-aminoacid Drosophila allatostatin receptor which is coupled to the Gi/Go class of G proteins.

The aggregation of the amyloid-␤ (A␤) peptide plays a pivotal role in the pathogenesis of Alzheimer's disease, as soluble oligomers are intimately linked to neuronal toxicity and inhibition of hippocampal long-term potentiation (LTP). In the C-terminal region of A␤ there are three consecutive GxxxG dimerization motifs, which we could previously demonstrate to play a critical role in the generation of A␤. Here, we show that glycine 33 (G33) of the central GxxxG interaction motif within the hydrophobic A␤ sequence is important for the aggregation dynamics of the peptide. A␤ peptides with alanine or isoleucine substitutions of G33 displayed an increased propensity to form higher oligomers, which we could attribute to conformational changes. Importantly, the oligomers of G33 variants were much less toxic than A␤ 42 wild type (WT), in vitro and in vivo. Also, whereas A␤ 42 WT is known to inhibit LTP, A␤ 42 G33 variants had lost the potential to inhibit LTP. Our findings reveal that conformational changes induced by G33 substitutions unlink toxicity and oligomerization of A␤ on the molecular level and suggest that G33 is the key amino acid in the toxic activity of A␤. Thus, a specific toxic conformation of A␤ exists, which represents a promising target for therapeutic interventions.