Cysteine oxidation and disulfide formation in the ribosomal exit tunnel.

Funder: DFG graduate college: CLiC State of Hesse HMWK: BMRZUnderstanding the conformational sampling of translation-arrested ribosome nascent chain complexes is key to understand co-translational folding. Up to now, coupling of cysteine oxidation, disulfide bond formation and structure formation in nascent chains has remained elusive. Here, we investigate the eye-lens protein γB-crystallin in the ribosomal exit tunnel. Using mass spectrometry, theoretical simulations, dynamic nuclear polarization-enhanced solid-state nuclear magnetic resonance and cryo-electron microscopy, we show that thiol groups of cysteine residues undergo S-glutathionylation and S-nitrosylation and form non-native disulfide bonds. Thus, covalent modification chemistry occurs already prior to nascent chain release as the ribosome exit tunnel provides sufficient space even for disulfide bond formation which can guide protein folding

Frankliniella occidentalis (Pergande) integrated pest management programs for fruiting vegetables in Florida

BACKGROUND: The spread of the western flower thrips, Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae), resulted in the worldwide destabilization of established integrated pest management programs for many crops. Efforts to control the pest and the thrips-vectored tospoviruses with calendar applications of broad-spectrum insecticides have been unsuccessful. The result has been a classic ‘3-R’ situation: resistance to numerous insecticides; resurgence of the western flower thrips populations as a result of natural predators and native competitor thrips being eliminated; replacement by various other pests. This paper reports on integrated pest management programs for fruiting vegetables that are effective, economical, ecologically sound and sustainable. RESULTS: The components include the following: define pest status (economic thresholds); increase biotic resistance (natural enemies and competition); integrate preventive and therapeutic tactics (scouting, ultraviolet-reflective technologies, biological control, compatible insecticides, companion plants and fertility); vertically integrate the programs with other pests; continually communicate latest science-based management tactics with end-users. CONCLUSION: These programs have been widely implemented in Florida and have significantly improved the management of western flower thrips and thrips-transmitted viruses.This is the publisher’s final pdf. The article is copyrighted by the Society of Chemical Industry and published by John Wiley & Sons, Inc. It can be found at: http://onlinelibrary.wiley.com/journal/10.1002/%28ISSN%291526-4998Keywords: thrips, fruiting vegetables, tospovirus, integrated pest management, Frankliniella occidentali

Non-Standard Errors

In statistics, samples are drawn from a population in a data-generating process (DGP). Standard errors measure the uncertainty in estimates of population parameters. In science, evidence is generated to test hypotheses in an evidence-generating process (EGP). We claim that EGP variation across researchers adds uncertainty: Non-standard errors (NSEs). We study NSEs by letting 164 teams test the same hypotheses on the same data. NSEs turn out to be sizable, but smaller for better reproducible or higher rated research. Adding peer-review stages reduces NSEs. We further find that this type of uncertainty is underestimated by participants

Untersuchungen zur Interaktion von Cholesterin mit cholesterinbindenden Proteinen

Der geschwindigkeitsbestimmende Schritt bei der Biosynthese von Steroidhormonen ist der Transport von Cholesterin von der äußeren zur inneren Mitochondrienmembran, wo es zu dem Steroid Pregnenolon umgewandelt wird. Für diesen Transport ist das StAR-Protein (Steroidogenic Acute Regulatory Protein) notwendig. Ein weiteres an der Bildung von Steroidhormonen beteiligtes Protein ist das MLN64-Protein. Beide Proteine besitzen so genannte START-Domänen (StAR related Lipid Transfer-Domänen), die Cholesterin binden können. In dieser Arbeit konnte gezeigt werden, dass die START-Domänen von StAR und MLN64 Cholesterin auf unterschiedliche Weise binden. Es ist noch nicht geklärt, auf welche Weise das StAR-Protein den Cholesterintransport in die Mitochondrien bewirkt. Das StAR-Protein könnte Cholesterin binden und als Cholesterintransporter zwischen äußerer und innerer Mitochondrienmembran fungieren. Nach einer anderen Hypothese wirkt das StAR-Protein ausschließlich an der äußeren Mitochondrienmembran. Es wird auch postuliert, dass das StAR-Protein in einem teilweise entfalteten Zustand vorliegen muss, um seine Funktion erfüllen zu können. In dieser Arbeit konnte gezeigt werden, dass StAR ein fotoreaktives Cholesterinderivat bindet. Die Cholesterinbindungsstelle des StAR-Proteins konnte eingegrenzt werden. Es wurden Experimente durchgeführt, um zu überprüfen, ob das Protein tatsächlich nur in teilweise entfaltetem Zustand aktiv ist. Die Cholesterinbindung des MLN64-Proteins wurde ebenfalls mit dem fotoreaktiven Cholesterinderivat untersucht. Dabei zeigte sich, dass MLN64 offenbar mehrere Bindungsstellen für Cholesterin besitzt. Weitere Experimente beschäftigten sich mit der Charakterisierung der Cholesterinbindungsstelle des humanen Oxytocinrezeptors, eines G-Protein gekoppelten Hormonrezeptors, der durch Cholesterin reguliert wird. Dabei kam auch wieder das fotoreaktive Cholesterinderivat zum Einsatz. Außerdem wurden in dieser Arbeit Experimente durchgeführt, die sich mit der Regulation der Cholesterinbiosynthese befassten. Die Biosynthese des Cholesterins wird reguliert, indem in der Membran des Endoplasmatischen Retikulums verankerte Transkriptionsfaktoren proteolytisch freigesetzt werden. Das passiert nur dann, wenn der zelluläre Cholesterinspiegel niedrig ist. Bei diesem Regulationsmechanismus spielt das Protein SCAP eine zentrale Rolle (Sterol responsive element binding protein Cleavage Activating Protein). SCAP bindet Cholesterin spezifisch und wird dadurch reguliert. Im Rahmen dieser Arbeit konnte der Bereich von SCAP eingegrenzt werden, der Cholesterin bindet. Ebenso konnte gezeigt werden, dass die Interaktion von SCAP mit einem anderen, als Insig bezeichneten Protein indirekt durch das Cholesterinderivat 25-Hydroxycholesterin reguliert wird.The rate limiting step in the acute biogenesis of steroid hormones is the transfer of cholesterol from the outer to the inner mitochondrial membrane where cholesterol is converted to pregnenolone. The StAR-protein (Steroidogenic Acute Regulatory Protein) is essential for the cholesterol transport to the inner mitochondrial membrane. MLN64 is another protein involved in the biogenesis of steroid hormones. Both proteins, StAR and MLN64, contain START-domains (StAR related Lipid Transfer-domains) which can bind cholesterol. In this work, it was shown that the START-domains of StAR and MLN64 bind cholesterol in different ways. It is not known yet how StAR enables the cholesterol transport into mitochondria. StAR could bind cholesterol and act as cholesterol shuttle between outer and inner mitochondrial membrane. According to another hypothesis, StAR works on the outer mitochondrial membrane. It was postulated that StAR must be partially unfolded to bind cholesterol. In this work it was shown that StAR binds a photoreactive cholesterol-analogue. The region of StAR which binds the cholesterol-analogue was identified. Experiments were performed to test the hypothesis if StAR must necessarily be partially unfolded to bind cholesterol. The interaction of MLN64 with cholesterol was examined. It was revealed that MLN64 -in opposite to StAR- must possess more than a single cholesterol binding site. Other Experiments had the aim to characterize the cholesterol binding site of the human Oxytocin-receptor which is regulated in its function by specific binding of cholesterol. The photoreactive cholesterol-analogue was used for these experiments. Furthermore, in this work, experiments were done to characterize some aspects of the regulation of cholesterol biosynthesis. The biosynthesis of cholesterol is regulated by proteoytical release of transcription factors from the membrane of the endoplasmic reticulum. The proteoytical cleavage is only possible if the cellular cholesterol level is low. In this regulation process, SCAP (Sterol responsive element binding protein Cleavage Activating Protein) plays an important role. SCAP binds cholesterol specifically and is regulated in its function by cholesterol-binding. In this work a region of SCAP could be identified which binds the photoreactive cholesterol-analogue. It was also shown that the interaction of SCAP with a protein called Insig is regulated in a indirect way by 25-Hydroxcholesterol

\u3ci\u3eFrankliniella occidentalis\u3c/i\u3e (Pergande) integrated pest management programs for fruiting vegetables in Florida

BACKGROUND:The spread of the western flower thrips, Frankliniella occidentalis (Pergande) (Thysanoptera: Thripidae), resulted in the worldwide destabilization of established integrated pest management programs for many crops. Efforts to control the pest and the thrips-vectored tospoviruses with calendar applications of broad-spectrum insecticides have been unsuccessful. The result has been a classic ‘3-R’ situation: resistance to numerous insecticides; resurgence of the western flower thrips populations as a result of natural predators and native competitor thrips being eliminated; replacement by various other pests. This paper reports on integrated pest management programs for fruiting vegetables that are effective, economical, ecologically sound and sustainable. RESULTS: The components include the following: define pest status (economic thresholds); increase biotic resistance (natural enemies and competition); integrate preventive and therapeutic tactics (scouting, ultraviolet-reflective technologies, biological control, compatible insecticides, companion plants and fertility); vertically integrate the programs with other pests; continually communicate latest science-based management tactics with end-users. CONCLUSION: These programs have been widely implemented in Florida and have significantly improved the management of western flower thrips and thrips-transmitted viruses

Cardiovascular disease and cognitive dysfunction in systemic lupus erythematosus

OBJECTIVE: Cognitive dysfunction and cardiovascular disease are common and debilitating manifestations of systemic lupus erythematosus (SLE). In this study, we evaluated the relationship between cardiovascular events, traditional cardiovascular risk factors, and SLE-specific risk factors as predictors of cognitive dysfunction in a large cohort of participants with SLE. METHODS: Subjects included 694 participants from the Lupus Outcomes Study (LOS), a longitudinal study of SLE outcomes based on annual telephone survey querying demographic and clinical variables. The Hopkins Verbal Learning Test – Revised (HVLT-R) and the Controlled Oral Word Association Test (COWAT) were administered to assess cognitive function. Multiple logistic regression was used to identify cardiovascular events (myocardial infarction (MI), stroke), traditional cardiovascular risk factors (hypertension, hyperlipidemia, diabetes, obesity, smoking), and SLE-specific risk factors (antiphospholipid antibodies (aPL), disease activity, disease duration) associated with cognitive impairment in year seven of the LOS. RESULTS: The prevalence of cognitive impairment as measured by verbal memory and verbal fluency metrics was 15%. In adjusted multiple logistic regression analyses, aPL (OR=2.10, 95% CI 1.3-3.41), hypertension (OR=2.06, 95% CI 1.19-3.56), and a history of stroke (OR=2.27, 95% CI 1.16-4.43) were significantly associated with cognitive dysfunction. In additional analyses evaluating the association between these predictors and severity of cognitive impairment, stroke was significantly more prevalent in participants with severe impairment when compared to those with mild or moderate impairment (p=0.036). CONCLUSIONS: These results suggest that the presence of aPL, hypertension, and stroke are key variables associated with cognitive impairment, which may aid in identification of patients at greatest risk

Structural and functional analysis of the promiscuous AcrB and AdeB efflux pumps suggests different drug binding mechanisms

Upon antibiotic stress Gram-negative pathogens deploy resistance-nodulation-cell division-type tripartite efflux pumps. These include a H+/drug antiporter module that recognizes structurally diverse substances, including antibiotics. Here, we show the 3.5 Å structure of subunit AdeB from the Acinetobacter baumannii AdeABC efflux pump solved by single-particle cryo-electron microscopy. The AdeB trimer adopts mainly a resting state with all protomers in a conformation devoid of transport channels or antibiotic binding sites. However, 10% of the protomers adopt a state where three transport channels lead to the closed substrate (deep) binding pocket. A comparison between drug binding of AdeB and Escherichia coli AcrB is made via activity analysis of 20 AdeB variants, selected on basis of side chain interactions with antibiotics observed in the AcrB periplasmic domain X-ray co-structures with fusidic acid (2.3 Å), doxycycline (2.1 Å) and levofloxacin (2.7 Å). AdeABC, compared to AcrAB-TolC, confers higher resistance to E. coli towards polyaromatic compounds and lower resistance towards antibiotic compounds