Transcription-Translation Coupling in Bacteria.

In bacteria, transcription and translation take place in the same cellular compartment. Therefore, a messenger RNA can be translated as it is being transcribed, a process known as transcription-translation coupling. This process was already recognized at the dawn of molecular biology, yet the interplay between the two key players, the RNA polymerase and ribosome, remains elusive. Genetic data indicate that an RNA sequence can be translated shortly after it has been transcribed. The closer both processes are in time, the less accessible the RNA sequence is between the RNA polymerase and ribosome. This temporal coupling has important consequences for gene regulation. Biochemical and structural studies have detailed several complexes between the RNA polymerase and ribosome. The in vivo relevance of this physical coupling has not been formally demonstrated. We discuss how both temporal and physical coupling may mesh to produce the phenomenon we know as transcription-translation coupling.

Orphan lysosomal solute carrier MFSD1 facilitates highly selective dipeptide transport.

Orphan solute carrier (SLC) represents a group of membrane transporters whose exact functions and substrate specificities are not known. Elucidating the function and regulation of orphan SLC transporters is not only crucial for advancing our knowledge of cellular and molecular biology but can potentially lead to the development of new therapeutic strategies. Here, we provide evidence for the biological function of a ubiquitous orphan lysosomal SLC, the Major Facilitator Superfamily Domain-containing Protein 1 (MFSD1), which has remained phylogenetically unassigned. Targeted metabolomics revealed that dipeptides containing either lysine or arginine residues accumulate in lysosomes of cells lacking MFSD1. Whole-cell patch-clamp electrophysiological recordings of HEK293-cells expressing MFSD1 on the cell surface displayed transport affinities for positively charged dipeptides in the lower mM range, while dipeptides that carry a negative net charge were not transported. This was also true for single amino acids and tripeptides, which MFSD1 failed to transport. Our results identify MFSD1 as a highly selective lysosomal lysine/arginine/histidine-containing dipeptide exporter, which functions as a uniporter.

Structural basis of a two-step tRNA recognition mechanism for plastid glycyl-tRNA synthetase.

Two types of glycyl-tRNA synthetase (GlyRS) are known, the α2 and the α2ß2 GlyRSs. Both types of synthetase employ a class II catalytic domain to aminoacylate tRNAGly. In plastids and some bacteria, the α and ß subunits are fused and are designated as (αß)2 GlyRSs. While the tRNA recognition and aminoacylation mechanisms are well understood for α2 GlyRSs, little is known about the mechanisms for α2ß2/(αß)2 GlyRSs. Here we describe structures of the (αß)2 GlyRS from Oryza sativa chloroplast by itself and in complex with cognate tRNAGly. The set of structures reveals that the U-shaped ß half of the synthetase selects the tRNA in a two-step manner. In the first step, the synthetase engages the elbow and the anticodon base C35 of the tRNA. In the second step, the tRNA has rotated ∼9° toward the catalytic centre. The synthetase probes the tRNA for the presence of anticodon base C36 and discriminator base C73. This intricate mechanism enables the tRNA to access the active site of the synthetase from a direction opposite to that of most other class II synthetases.

Ang II Promotes ET-1 Production by Regulating NOX2 Activity Through Transcription Factor Oct-1.

BACKGROUND: Increasing evidence suggests that superoxide ions produced by NOX (nicotinamide adenine dinucleotide phosphate oxidases) mediate vascular effects of Ang II (angiotensin II) evoked by atherogenic diets. Here, we analyzed the mechanism by which NOX2 contributes to Ang II-induced ET-1 (endothelin 1) production in human microvascular endothelial cells. METHODS: The effects of high-fat diet were compared between WT (wild type) and Nox2 (mouse NOX2 gene)-deficient mice. ET-1 production and NOX2 expression by human microvascular endothelial cells in vitro were analyzed by ELISA, reverse transcription quantitative polymerase chain reaction, electrophoretic mobility shift assay, promoter deletions, RNA interference, and pharmacological inhibition. Production of superoxide anions was visualized by fluorescent cell labeling. RESULTS: Feeding mice high-fat diet for 10 weeks increased cardiac expression and plasma levels of Ang II and ET-1 in WT but not in Nox2-deficient animals. Exposure of human microvascular endothelial cells to Ang II resulted in increased ET-1 production, which could be blocked by silencing NOX2 (human NOX2 gene). Ang II promoted NOX2 expression through induction of the Oct-1 (human/mouse octamer binding transcription factor 1 protein) and activation of the NOX2 promoter region containing Oct-1-binding sites. Stimulation of NOX2 expression by Ang II was associated with increased production of superoxide anions. Inhibition of Oct-1 by small interfering RNA reduced Ang II-induced NOX2 expression and superoxide anion production, and neutralization of superoxide by SOD (superoxide dismutase) abolished Ang II-stimulated ET1 (human ET-1 gene) promoter activity, ET1 mRNA expression, and ET-1 release. CONCLUSIONS: Ang II may promote ET-1 production in the endothelium in response to atherogenic diets through a mechanism that involves the transcription factor Oct-1 and the increased formation of superoxide anions by NOX2.

Repeated Evolution of Unorthodox Feeding Styles Drives a Negative Correlation between Foot Size and Bill Length in Hummingbirds.

AbstractDifferences among hummingbird species in bill length and shape have rightly been viewed as adaptive in relation to the morphology of the flowers they visit for nectar. In this study we examine functional variation in a behaviorally related but neglected feature: hummingbird feet. We gathered records of hummingbirds clinging by their feet to feed legitimately as pollinators or illegitimately as nectar robbers-"unorthodox" feeding behaviors. We measured key features of bills and feet for 220 species of hummingbirds and compared the 66 known "clinger" species (covering virtually the entire scope of hummingbird body size) with the 144 presumed "non-clinger" species. Once the effects of phylogenetic signal, body size, and elevation above sea level are accounted for statistically, hummingbirds display a surprising but functionally interpretable negative correlation. Clingers with short bills and long hallux (hind-toe) claws have evolved-independently-more than 20 times and in every major clade. Their biomechanically enhanced feet allow them to save energy by clinging to feed legitimately on short-corolla flowers and by stealing nectar from long-corolla flowers. In contrast, long-billed species have shorter hallux claws, as plant species with long-corolla flowers enforce hovering to feed, simply by the way they present their flowers.

Chronic post-COVID-19 syndrome and chronic fatigue syndrome: Is there a role for extracorporeal apheresis?

As millions of patients have been infected by SARS-CoV-2 virus a vast number of individuals complain about continuing breathlessness and fatigue even months after the onset of the disease. This overwhelming phenomenon has not been well defined and has been called "post-COVID syndrome" or "long-COVID" [1]. There are striking similarities to myalgic encephalomyelitis also called chronic fatigue syndrome linked to a viral and autoimmune pathogenesis. In both disorders neurotransmitter receptor antibodies against ß-adrenergic and muscarinic receptors may play a key role. We found similar elevation of these autoantibodies in both patient groups. Extracorporeal apheresis using a special filter seems to be effective in reducing these antibodies in a significant way clearly improving the debilitating symptoms of patients with chronic fatigue syndrome. Therefore, such a form of neuropheresis may provide a promising therapeutic option for patients with post-COVID-19 syndrome. This method will also be effective when other hitherto unknown antibodies and inflammatory mediators are involved.

Crystallographic and kinetic analyses of the FdsBG subcomplex of the cytosolic formate dehydrogenase FdsABG from Cupriavidus necator.

Formate oxidation to carbon dioxide is a key reaction in one-carbon compound metabolism, and its reverse reaction represents the first step in carbon assimilation in the acetogenic and methanogenic branches of many anaerobic organisms. The molybdenum-containing dehydrogenase FdsABG is a soluble NAD+-dependent formate dehydrogenase and a member of the NADH dehydrogenase superfamily. Here, we present the first structure of the FdsBG subcomplex of the cytosolic FdsABG formate dehydrogenase from the hydrogen-oxidizing bacterium Cupriavidus necator H16 both with and without bound NADH. The structures revealed that the two iron-sulfur clusters, Fe4S4 in FdsB and Fe2S2 in FdsG, are closer to the FMN than they are in other NADH dehydrogenases. Rapid kinetic studies and EPR measurements of rapid freeze-quenched samples of the NADH reduction of FdsBG identified a neutral flavin semiquinone, FMNH•, not previously observed to participate in NADH-mediated reduction of the FdsABG holoenzyme. We found that this semiquinone forms through the transfer of one electron from the fully reduced FMNH-, initially formed via NADH-mediated reduction, to the Fe2S2 cluster. This Fe2S2 cluster is not part of the on-path chain of iron-sulfur clusters connecting the FMN of FdsB with the active-site molybdenum center of FdsA. According to the NADH-bound structure, the nicotinamide ring stacks onto the re-face of the FMN. However, NADH binding significantly reduced the electron density for the isoalloxazine ring of FMN and induced a conformational change in residues of the FMN-binding pocket that display peptide-bond flipping upon NAD+ binding in proper NADH dehydrogenases.

The Lysozyme Inhibitor Thionine Acetate Is Also an Inhibitor of the Soluble Lytic Transglycosylase Slt35 from Escherichia coli.

Lytic transglycosylases such as Slt35 from E. coli are enzymes involved in bacterial cell wall remodelling and recycling, which represent potential targets for novel antibacterial agents. Here, we investigated a series of known glycosidase inhibitors for their ability to inhibit Slt35. While glycosidase inhibitors such as 1-deoxynojirimycin, castanospermine, thiamet G and miglitol had no effect, the phenothiazinium dye thionine acetate was found to be a weak inhibitor. IC50 values and binding constants for thionine acetate were similar for Slt35 and the hen egg white lysozyme. Molecular docking simulations suggest that thionine binds to the active site of both Slt35 and lysozyme, although it does not make direct interactions with the side-chain of the catalytic Asp and Glu residues as might be expected based on other inhibitors. Thionine acetate also increased the potency of the beta-lactam antibiotic ampicillin against a laboratory strain of E. coli.

An amino-terminal threonine/serine motif is necessary for activity of the Crp/Fnr homolog, MrpC and for Myxococcus xanthus developmental robustness.

The Crp/Fnr family of transcriptional regulators play central roles in transcriptional control of diverse physiological responses, and are activated by a surprising diversity of mechanisms. MrpC is a Crp/Fnr homolog that controls the Myxococcus xanthus developmental program. A long-standing model proposed that MrpC activity is controlled by the Pkn8/Pkn14 serine/threonine kinase cascade, which phosphorylates MrpC on threonine residue(s) located in its extreme amino-terminus. In this study, we demonstrate that a stretch of consecutive threonine and serine residues, T21 T22 S23 S24, is necessary for MrpC activity by promoting efficient DNA binding. Mass spectrometry analysis indicated the TTSS motif is not directly phosphorylated by Pkn14 in vitro but is necessary for efficient Pkn14-dependent phosphorylation on several residues in the remainder of the protein. In an important correction to a long-standing model, we show Pkn8 and Pkn14 kinase activities do not play obvious roles in controlling MrpC activity in wild-type M. xanthus under laboratory conditions. Instead, we propose Pkn14 modulates MrpC DNA binding in response to unknown environmental conditions. Interestingly, substitutions in the TTSS motif caused developmental defects that varied between biological replicates, revealing that MrpC plays a role in promoting a robust developmental phenotype.

Transcription-translation coupling: direct interactions of RNA polymerase with ribosomes and ribosomal subunits.

In prokaryotes, RNA polymerase and ribosomes can bind concurrently to the same RNA transcript, leading to the functional coupling of transcription and translation. The interactions between RNA polymerase and ribosomes are crucial for the coordination of transcription with translation. Here, we report that RNA polymerase directly binds ribosomes and isolated large and small ribosomal subunits. RNA polymerase and ribosomes form a one-to-one complex with a micromolar dissociation constant. The formation of the complex is modulated by the conformational and functional states of RNA polymerase and the ribosome. The binding interface on the large ribosomal subunit is buried by the small subunit during protein synthesis, whereas that on the small subunit remains solvent-accessible. The RNA polymerase binding site on the ribosome includes that of the isolated small ribosomal subunit. This direct interaction between RNA polymerase and ribosomes may contribute to the coupling of transcription to translation.

Two Old Dogs, One New Trick: A Review of RNA Polymerase and Ribosome Interactions during Transcription-Translation Coupling.

The coupling of transcription and translation is more than mere translation of an mRNA that is still being transcribed. The discovery of physical interactions between RNA polymerase and ribosomes has spurred renewed interest into this long-standing paradigm of bacterial molecular biology. Here, we provide a concise presentation of recent insights gained from super-resolution microscopy, biochemical, and structural work, including cryo-EM studies. Based on the presented data, we put forward a dynamic model for the interaction between RNA polymerase and ribosomes, in which the interactions are repeatedly formed and broken. Furthermore, we propose that long intervening nascent RNA will loop out and away during the forming the interactions between the RNA polymerase and ribosomes. By comparing the effect of the direct interactions between RNA polymerase and ribosomes with those that transcription factors NusG and RfaH mediate, we submit that two distinct modes of coupling exist: Factor-free and factor-mediated coupling. Finally, we provide a possible framework for transcription-translation coupling and elude to some open questions in the field.

Age and growth of three endemic threatened guitarfishes Pseudobatos horkelii, P. percellens and Zapteryx brevirostris in the western South Atlantic Ocean.

The age and growth of three endemic threatened guitarfish species were analysed using vertebrae of Pseudobatos horkelii, P. percellens and Zapteryx brevirostris. Edge and marginal-increment analyses were used to evaluate the periodicity of the formation of the band-pairs, suggesting deposition of one band-pair per year, from late winter to late spring. The von Bertalanffy growth model was used to describe the growth of these species with the following parameters, for pooled sexes: P. horkelii L∞ = 126.93, k = 0.19 and t0 = -1.51; P. percellens L∞ = 109.31, k = 0.16 and t0 = -1.78; Z. brevirostris L∞ = 60.37, k = 0.24 and t0 = -1.42. Our results are essential to understanding the resilience and vulnerability of these species to harvest, which can contribute to management and conservation actions of these species.

Erratum: Measurement of Body-Centered-Cubic Aluminum at 475 GPa [Phys. Rev. Lett. 119, 175702 (2017)].

This corrects the article DOI: 10.1103/PhysRevLett.119.175702.

Use of whole body CT to detect patterns of CPR-related injuries after sudden cardiac arrest.

AIMS AND OBJECTIVES: We have recently implemented a dedicated sudden cardiac arrest (SCA) - whole-body computed tomography (WBCT) protocol to evaluate SCA patients with return of spontaneous circulation (ROSC) following cardiopulmonary resuscitation (CPR). The aim of this study is to evaluate the number and pattern of CPR-related injuries in ROSC patients with SCA-WBCT. METHODS AND MATERIALS: Single-centre retrospective review of 39 patients (13 female; 20 male, mean age 51.8 years) with non-traumatic, out-of-hospital SCA and ROSC and evaluation with dedicated SCA-WBCT over a 10-month period. RESULTS: In-hospital mortality was 54%. CPR-related injuries were detected in 85% (33/39). Chest injuries were most common on WBCT: 85% (33) subjects had rib fractures (mean of 8.5 fractures/subject); 31% (12) sternal fractures; 13% (5) mediastinal haematoma; 10% (4) pneumothorax; 8% (3) pneumomediastinum and 3% (1) haemothorax. Three subjects (8%) had abdominal injuries on WBCT, including one hepatic haematoma with active haemorrhage. CONCLUSION: CPR-related injuries on WBCT after ROSC are common, with serial rib fractures detected most commonly. An unexpectedly high rate of abdominal injuries was detected on SCA-WBCT. Radiologists need to be attuned to the spectrum of CPR-related injuries in WBCT, including abdominal injuries and subtle rib fractures. KEY POINTS: • CPR frequently causes injuries. • Radiologists should be aware of the spectrum of CPR related injuries. • Rib fractures are frequent and radiologic findings often subtle. • Clinically unexpected abdominal injuries may be present.

In-hospital usability and feasibility evaluation of Panda, an app for the management of pain in children at home.

BACKGROUND: Postoperative pain in children is often poorly managed at home, leading to slower functional recovery, poor oral intake, sleep disturbances, and behavioral changes. Panda is a smartphone application (app) designed to support parents in assessing their child's pain and managing medications. AIMS: The aim of this study was to evaluate the Panda app's usability and feasibility in hospital prior to testing the app at home. METHODS: The study comprised two phases. Phase I evaluated Panda's usability with nurses, parents, and adolescents using simulated scenarios. Usability was measured by task completion rate, user error rates, and the Computer Systems Usability Questionnaire. Phase II evaluated Panda's feasibility by observing parents/guardians of pediatric patients using the app on the postsurgical ward. Feasibility was measured using response frequency and delay following app notifications from an audit trail of app function, and parental satisfaction from an interview. Feedback was used to guide iterative app improvements. RESULTS: In Phase I, 13 nurses, 12 parents, and 5 adolescents evaluated the app. A total of 103 usability issues were identified, analyzed, and addressed. In Phase II, 29 parents responded to a total of 151 app notifications, with 84% responding within 1 hour in the final round of testing; 93% of participants reported the app was easy to use, and rated the app with a median [interquartile range] Computer Systems Usability Questionnaire score of 2 [1-4]. Significant barriers to use included lack of flexibility in the medication scheduling, low volume of alert sounds, and the extra time spent on medication safety checks. CONCLUSION: Panda's usability was improved and its feasibility demonstrated in the controlled hospital environment. The next step is to evaluate its feasibility for use at home.

Systematic Review of Quality-of-Life Measurement After Aesthetic Rhinoplasty.

INTRODUCTION: The assessment of outcomes in aesthetic rhinoplasty is highly relevant because patient satisfaction and improved health-related quality of life (QoL) are the predominant factors in determining success. The patient-reported outcome measures (PROMs) employed in rhinoplasty research studies are remarkably diverse, thus yielding difficulties with data analysis. The aim of this article is to provide a comprehensive review of the literature to reveal the relevance of the QoL assessment for rhinosurgeons. METHODS: A systematic literature search with the terms "Rhinoplasty" and "Quality of Life" was conducted using PubMed/MEDLINE, Google Scholar and Cochrane databases. Primarily, all publications related to QoL following aesthetic rhinoplasty between 2002 and 2017 were identified. As a secondary selection, we focused on articles with a prospective study design, a significant cohort size (at least 50 patients) and a follow-up period of at least 6 months after aesthetic rhinoplasty. RESULTS: A total of 62 PROM studies assessing QoL following aesthetic rhinoplasties were obtained. We ascertained an increasing publication rate of QoL articles over the last 15 years. Only 17 studies satisfied comprehensive inclusion criteria of a high qualitative study selection. The Rhinoplasty Outcome Evaluation was the most frequently used QoL questionnaire of the secondary selection (70.6%). The total number of 16 various questionnaires exhibit high heterogeneity. CONCLUSION: Our data strengthen the increasing importance of the assessment of QoL after rhinoplasty. Despite a lack of reliable publications with considerable heterogeneity and large variability in outcomes, functional-aesthetic rhinoplasty leads to a significant improvement of patient's health-related QoL. LEVEL OF EVIDENCE II: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266 .

Measurement of Body-Centered-Cubic Aluminum at 475 GPa.

Nanosecond in situ x-ray diffraction and simultaneous velocimetry measurements were used to determine the crystal structure and pressure, respectively, of ramp-compressed aluminum at stress states between 111 and 475 GPa. The solid-solid Al phase transformations, fcc-hcp and hcp-bcc, are observed at 216±9 and 321±12 GPa, respectively, with the bcc phase persisting to 475 GPa. The high-pressure crystallographic texture of the hcp and bcc phases suggests close-packed or nearly close-packed lattice planes remain parallel through both transformations.

ASER core curriculum illustration project: aortic intramural hematoma (IMH).

This is the 23rd installment of a series that will highlight one case per publication issue from the bank of cases available online as part of the American Society of Emergency Radiology (ASER) educational resources. Our goal is to generate more interest in and use of our online materials. To view more cases online, please visit the ASER Core Curriculum and Recommendations for Study online at http://www.aseronline.org/curriculum/toc.htm .

Core curriculum illustration: rib fractures.

This is the 24th installment of a series that will highlight one case per publication issue from the bank of cases available online as part of the American Society of Emergency Radiology (ASER) educational resources. Our goal is to generate more interest in and use of our online materials. To view more cases online, please visit the ASER Core Curriculum and Recommendations for Study online at http://www.aseronline.org/curriculum/toc.htm .