Risk-specific search for risk-defusing operators

According to the concept of “active risk-defusing behavior”, decision makers in risky situations look for additional actions that reduce risk and allow them to favor the more risky alternative. Our study demonstrates that risk-defusing behavior depends on the type of risk (normal, medium, catastrophic or global) as well as on the domain (health, economy or ecology). In total, 12 scenarios (four risk types from three risk domains each) were constructed. Using the interview techniques of active information search and thinking-aloud, 120 interviews about decision-making processes with these scenarios were conducted. They showed that the active search for different risk-defusing operators depends on the type of risk, but even more on the domain of the scenario. Results suggest a need for further research about a typology of risk situations in which, besides formal classification criteria, content issues are also explored

Role of bacterial isolates in enhancing the bud induction in the industrially important red alga Gracilaria dura

Plant growth depends on the integration of environmental cues, nitrogen fixation and phytohormone-signaling pathways. The growth and development of Gracilaria dura was significantly influenced by the association of bacterial isolates. The putative bud-inducing epiphytic Exiguobacterium homiense and endophytic Bacillus pumilus, Bacillus licheniformis were examined for their ability to fix nitrogen and produce indole-3-acetic acid (IAA). These bacterial isolates were identified to the species level by biochemical tests, fatty acid and partial 16S rRNA gene sequence analysis. The B. pumilus, B. licheniformis and E. homiense produced 445.5, 335 and 184.1 μg mL−1 IAA and 12.51, 10.14 and 6.9 mM mL−1 ammonium, respectively, as determined using HPLC and spectroscopy. New bud regeneration observed after the addition of total protein of the bacterial isolates suggests that IAA is conjugated with protein. The epi- and endophytic bacterial isolates were able to induce five and 10 new buds per frond, respectively, in comparison to the control, where one to two buds were observed. The combination of 25 °C and 30‰ showed the optimum condition for bud induction in G. dura when incubated with the total protein of B. pumilus. Our finding revealed for the first time that IAA coupled with nitrogen fixation induce and regenerate new buds in G. dur

Construction and characterization of Enterococcus faecalis CG110/gfp/pRE25*, a tool for monitoring horizontal gene transfer in complex microbial ecosystems

Enterococci are among the most notorious bacteria involved in the spread of antibiotic resistance (ABR) determinants via horizontal gene transfer, a process that leads to increased prevalence of antibiotic-resistant bacteria. In complex microbial communities with a high background of ABR genes, detection of gene transfer is possible only when the ABR determinant is marked. Therefore, the conjugative multiresistance plasmid pRE25, originating from a sausage-associated Enterococcus faecalis, was tagged with a 34-bp random sequence marker spliced by tet(M). The plasmid constructed, designated pRE25*, was introduced into E. faecalis CG110/gfp, a strain containing a gfp gene as chromosomal marker. The plasmid pRE25* is fully functional compared with its parental pRE25, occurs at one to two copies per chromosome, and can be transferred to Listeria monocytogenes and Listeria innocua at frequencies of 6 × 10−6 to 8 × 10−8 transconjugants per donor. The markers on the chromosome and the plasmid enable independent quantification of donor and plasmid, even if ABR genes occur at high numbers in the background ecosystem. Both markers were stable for at least 200 generations, permitting application of the strain in long-running experiments. Enterococcus faecalis CG110/gfp/pRE25* is a potent tool for the investigation of horizontal ABR gene transfer in complex environments such as food matrices, biofilms or colonic model

Fangschreckenkrebse – Superlative in der Tierwelt: Präparation eines Gliederfüßers zur Erarbeitung von Struktur-Funktionszusammenhängen

Superlative begeistern Kinder und Jugendliche. Fangschreckenkrebse (Stomatopoden) zeigen einige Merkmale, die man als „Superlative“ bezeichnen könnte. Zunächst fallen dabei die Fangmechanismen der Stomatopoden auf. Man unterscheidet gemein gebräuchlich sogenannte “Speerer” und “Boxer”. Die “Speerer” erreichen mit ihrem großen Raubbein eine Geschwindigkeit von über 100 km/h beim Beutefang, eine der schnellsten muskelvermittelten Bewegungen in der Tierwelt. Die “Boxer“ erzeugen eine Krafteinwirkung ähnlich einem Wolfsbiss oder einer Kleinkaliberkugel mit darauffolgender Druckwelle zum Zertrümmern der Schalen von Beutetieren. Außerdem ist der Besitz eines der besonderen optischen Apparate mit den meisten Fotorezeptortypen im gesamten Tierreich beeindruckend (Cronin et al. 2017).  Doch nicht nur diese Superlative machen die Fangschreckenkrebse für den Unterricht interessant. Fangschreckenkrebse sind leicht und kostengünstig zu besorgen und  mit einer Verkaufsgröße von bis zu ca. 25 cm sind die Extremitäten gut und anschaulich zu präparieren.  Die Extremitäten weisen eine morphologische Vielfalt auf, die als Angepasstheiten an die unterschiedlichen Funktionen zu verstehen sind.   So lassen sich die vielfältigen Variationen einer grundlegenden Körperorganisation am Original zeigen

Non-canonical shedding of TNFα by SPPL2a is determined by the conformational flexibility of its transmembrane helix

Ectodomain (EC) shedding defines the proteolytic removal of a membrane protein EC and acts as an important molecular switch in signaling and other cellular processes. Using tumor necrosis factor (TNF)α as a model substrate, we identify a non-canonical shedding activity of SPPL2a, an intramembrane cleaving aspartyl protease of the GxGD type. Proline insertions in the TNFα transmembrane (TM) helix strongly increased SPPL2a non-canonical shedding, while leucine mutations decreased this cleavage. Using biophysical and structural analysis, as well as molecular dynamic simulations, we identified a flexible region in the center of the TNFα wildtype TM domain, which plays an important role in the processing of TNFα by SPPL2a. This study combines molecular biology, biochemistry, and biophysics to provide insights into the dynamic architecture of a substrate\u27s TM helix and its impact on non-canonical shedding. Thus, these data will provide the basis to identify further physiological substrates of non-canonical shedding in the future

Non-canonical Shedding of TNFα by SPPL2a Is Determined by the Conformational Flexibility of Its Transmembrane Helix

Ectodomain (EC) shedding defines the proteolytic removal of a membrane protein EC and acts as an important molecular switch in signaling and other cellular processes. Using tumor necrosis factor (TNF)α as a model substrate, we identify a non-canonical shedding activity of SPPL2a, an intramembrane cleaving aspartyl protease of the GxGD type. Proline insertions in the TNFα transmembrane (TM) helix strongly increased SPPL2a non-canonical shedding, while leucine mutations decreased this cleavage. Using biophysical and structural analysis, as well as molecular dynamic simulations, we identified a flexible region in the center of the TNFα wildtype TM domain, which plays an important role in the processing of TNFα by SPPL2a. This study combines molecular biology, biochemistry, and biophysics to provide insights into the dynamic architecture of a substrate\u27s TM helix and its impact on non-canonical shedding. Thus, these data will provide the basis to identify further physiological substrates of non-canonical shedding in the future

Helical stability of the GnTV transmembrane domain impacts on SPPL3 dependent cleavage

Signal-Peptide Peptidase Like-3 (SPPL3) is an intramembrane cleaving aspartyl protease that causes secretion of extracellular domains from type-II transmembrane proteins. Numerous Golgi-localized glycosidases and glucosyltransferases have been identified as physiological SPPL3 substrates. By SPPL3 dependent processing, glycan-transferring enzymes are deactivated inside the cell, as their active site-containing domain is cleaved and secreted. Thus, SPPL3 impacts on glycan patterns of many cellular and secreted proteins and can regulate protein glycosylation. However, the characteristics that make a substrate a favourable candidate for SPPL3-dependent cleavage remain unknown. To gain insights into substrate requirements, we investigated the function of a GxxxG motif located in the transmembrane domain of N-acetylglucosaminyltransferase V (GnTV), a well-known SPPL3 substrate. SPPL3-dependent secretion of the substrate’s ectodomain was affected by mutations disrupting the GxxxG motif. Using deuterium/hydrogen exchange and NMR spectroscopy, we studied the effect of these mutations on the helix flexibility of the GnTV transmembrane domain and observed that increased flexibility facilitates SPPL3-dependent shedding and vice versa. This study provides first insights into the characteristics of SPPL3 substrates, combining molecular biology, biochemistry, and biophysical techniques and its results will provide the basis for better understanding the characteristics of SPPL3 substrates with implications for the substrates of other intramembrane proteases

Helical stability of the GnTV transmembrane domain impacts on SPPL3 dependent cleavage

Signal-Peptide Peptidase Like-3 (SPPL3) is an intramembrane cleaving aspartyl protease that causes secretion of extracellular domains from type-II transmembrane proteins. Numerous Golgi-localized glycosidases and glucosyltransferases have been identified as physiological SPPL3 substrates. By SPPL3 dependent processing, glycan-transferring enzymes are deactivated inside the cell, as their active site-containing domain is cleaved and secreted. Thus, SPPL3 impacts on glycan patterns of many cellular and secreted proteins and can regulate protein glycosylation. However, the characteristics that make a substrate a favourable candidate for SPPL3-dependent cleavage remain unknown. To gain insights into substrate requirements, we investigated the function of a GxxxG motif located in the transmembrane domain of N-acetylglucosaminyltransferase V (GnTV), a well-known SPPL3 substrate. SPPL3-dependent secretion of the substrate’s ectodomain was affected by mutations disrupting the GxxxG motif. Using deuterium/hydrogen exchange and NMR spectroscopy, we studied the effect of these mutations on the helix flexibility of the GnTV transmembrane domain and observed that increased flexibility facilitates SPPL3-dependent shedding and vice versa. This study provides first insights into the characteristics of SPPL3 substrates, combining molecular biology, biochemistry, and biophysical techniques and its results will provide the basis for better understanding the characteristics of SPPL3 substrates with implications for the substrates of other intramembrane proteases

Signal Peptide Peptidase-Like 2c (SPPL2c) impairs vesicular transport and cleavage of SNARE proteins

Members of the GxGD-type intramembrane aspartyl proteases have emerged as key players not only in fundamental cellular processes such as B-cell development or protein glycosylation, but also in development of pathologies, such as Alzheimer's disease or hepatitis virus infections. However, one member of this protease family, signal peptide peptidase-like 2c (SPPL2c), remains orphan and its capability of proteolysis as well as its physiological function is still enigmatic. Here, we demonstrate that SPPL2c is catalytically active and identify a variety of SPPL2c candidate substrates using proteomics. The majority of the SPPL2c candidate substrates cluster to the biological process of vesicular trafficking. Analysis of selected SNARE proteins reveals proteolytic processing by SPPL2c that impairs vesicular transport and causes retention of cargo proteins in the endoplasmic reticulum. As a consequence, the integrity of subcellular compartments, in particular the Golgi, is disturbed. Together with a strikingly high physiological SPPL2c expression in testis, our data suggest involvement of SPPL2c in acrosome formation during spermatogenesis