Structural basis for RNA-genome recognition during bacteriophage Qβ replication

Upon infection of Escherichia coli by bacteriophage Qβ, the virus-encoded β-subunit recruits host translation elongation factors EF-Tu and EF-Ts and ribosomal protein S1 to form the Qβ replicase holoenzyme complex, which is responsible for amplifying the Qβ (+)-RNA genome. Here, we use X-ray crystallography, NMR spectroscopy, as well as sequence conservation, surface electrostatic potential and mutational analyses to decipher the roles of the β-subunit and the first two oligonucleotide-oligosaccharide-binding domains of S1 (OB_1–2) in the recognition of Qβ (+)-RNA by the Qβ replicase complex. We show how three basic residues of the β subunit form a patch located adjacent to the OB₂ domain, and use NMR spectroscopy to demonstrate for the first time that OB₂ is able to interact with RNA. Neutralization of the basic residues by mutagenesis results in a loss of both the phage infectivity in vivo and the ability of Qβ replicase to amplify the genomic RNA in vitro. In contrast, replication of smaller replicable RNAs is not affected. Taken together, our data suggest that the β-subunit and protein S1 cooperatively bind the (+)-stranded Qβ genome during replication initiation and provide a foundation for understanding template discrimination during replication initiation.     

Plant defence as a complex and changing phenotype throughout ontogeny

Background and Aims Ontogenetic changes in anti-herbivore defences are common and result from variation in resource availability and herbivore damage throughout plant development. However, little is known about the simultaneous changes of multiple defences across the entire development of plants, and how such changes affect plant damage in the field. The aim of this study was to assess if changes in the major types of plant resistance and tolerance can explain natural herbivore damage throughout plant ontogeny.Methods An assessment was made of how six defensive traits, including physical, chemical and biotic resistance, simultaneously change across the major transitions of plant development, from seedlings to reproductive stages of Turnera velutina growing in the greenhouse. In addition, an experiment was performed to assess how plant tolerance to artificial damage to leaves changed throughout ontogeny. Finally, leaf damage by herbivores was evaluated in a natural population.Key Results The observed ontogenetic trajectories of all defences were significantly different, sometimes showing opposite directions of change. Whereas trichome density, leaf toughness, extrafloral nectary abundance and nectar production increased, hydrogen cyanide and compensatory responses decreased throughout plant development, from seedlings to reproductive plants. Only water content was higher at the intermediate juvenile ontogenetic stages. Surveys in a natural population over 3 years showed that herbivores consumed more tissue from juvenile plants than from younger seedlings or older reproductive plants. This is consistent with the fact that juvenile plants were the least defended stage.Conclusions The results suggest that defensive trajectories are a mixed result of predictions by the Optimal Defence Theory and the Growth–Differentiation Balance Hypothesis. The study emphasizes the importance of incorporating multiple defences and plant ontogeny into further studies for a more comprehensive understanding of plant defence evolution.     

A simple approximation for larval retention around reefs

Estimating larval retention at individual reefs by local scale three-dimensional flows is a significant problem for understanding, and predicting, larval dispersal. Determining larval dispersal commonly involves the use of computationally demanding and expensively calibrated/validated hydrodynamic models that resolve reef wake eddies. This study models variation in larval retention times for a range of reef shapes and circulation regimes, using a reef-scale three-dimensional hydrodynamic model. It also explores how well larval retention time can be estimated based on the “Island Wake Parameter”, a measure of the degree of flow turbulence in the wake of reefs that is a simple function of flow speed, reef dimension, and vertical diffusion. The mean residence times found in the present study (0.48–5.64 days) indicate substantial potential for self-recruitment of species whose larvae are passive, or weak swimmers, for the first several days after release. Results also reveal strong and significant relationships between the Island Wake Parameter and mean residence time, explaining 81–92% of the variability in retention among reefs across a range of unidirectional flow speeds and tidal regimes. These findings suggest that good estimates of larval retention may be obtained from relatively coarse-scale characteristics of the flow, and basic features of reef geomorphology. Such approximations may be a valuable tool for modeling connectivity and meta-population dynamics over large spatial scales, where explicitly characterizing fine-scale flows around reef requires a prohibitive amount of computation and extensive model calibration.     

Excitons in Metal‐Halide Perovskites

The unprecedented increase of the power conversion efficiency of metal‐halide perovskite solar cells has significantly outpaced the understanding of their fundamental properties. One of the biggest puzzles of perovskites has been the exciton binding energy, which has proved to be difficult to determine experimentally. Many contradictory reports can be found in the literature with values of the exciton binding energy from a few meV to a few tens of meV. In this review the results of the last few years of intense investigation of the exciton physic in perovskite materials are summarized. In particular a critical overview of the different experimental approaches used to determine exciton binding energy is provided. The problem of exciton binding energy in the context of the polar nature of perovskite crystals and related polaron effects which have been neglected to date in most of work is discussed. It is shown that polaron effects can reconcile at least some of the experimental observations and controversy present in the literature. Finally, the current status of the exciton fine structure in perovskite materials is summarized. The peculiar carrier–phonon coupling can help to understand the intriguing efficiency of light emission from metal‐halide perovskites.     

NETosis occurs independently of neutrophil serine proteases

Neutrophils are primary host innate immune cells defending against pathogens. One proposed mechanism by which neutrophils prevent the spread of pathogens is NETosis, the extrusion of cellular DNA resulting in neutrophil extracellular traps (NETs). The protease neutrophil elastase (NE) has been implicated in the formation of NETs through proteolysis of nuclear proteins leading to chromatin decondensation. In addition to NE, neutrophils contain three other serine proteases that could compensate if the activity of NE was neutralized. However, whether they do play such a role is unknown. Thus, we deployed recently described specific inhibitors against all four of the neutrophil serine proteases (NSPs). Using specific antibodies to the NSPs along with our labeled inhibitors, we show that catalytic activity of these enzymes is not required for the formation of NETs. Moreover, the NSPs that decorate NETs are in an inactive conformation and thus cannot participate in further catalytic events. These results indicate that NSPs play no role in either NETosis or arming NETs with proteolytic activity.     

Biosynthesis of GlcNAc-rich N- and O-glycans in the Golgi apparatus does not require the nucleotide sugar transporter SLC35A3

Nucleotide sugar transporters, encoded by the SLC35 gene family, deliver nucleotide sugars throughout the cell for various glycosyltransferase-catalyzed glycosylation reactions. GlcNAc, in the form of UDP-GlcNAc, and galactose, as UDP-Gal, are delivered into the Golgi apparatus by SLC35A3 and SLC35A2 transporters, respectively. However, although the UDP-Gal transporting activity of SLC35A2 has been clearly demonstrated, UDP-GlcNAc delivery by SLC35A3 is not fully understood. Therefore, we analyzed a panel of CHO, HEK293T, and HepG2 cell lines including WT cells, SLC35A2 knockouts, SLC35A3 knockouts, and double-knockout cells. Cells lacking SLC35A2 displayed significant changes in N- and O-glycan synthesis. However, in SLC35A3-knockout CHO cells, only limited changes were observed; GlcNAc was still incorporated into N-glycans, but complex type N-glycan branching was impaired, although UDP-GlcNAc transport into Golgi vesicles was not decreased. In SLC35A3-knockout HEK293T cells, UDP-GlcNAc transport was significantly decreased but not completely abolished. However, N-glycan branching was not impaired in these cells. In CHO and HEK293T cells, the effect of SLC35A3 deficiency on N-glycan branching was potentiated in the absence of SLC35A2. Moreover, in SLC35A3-knockout HEK293T and HepG2 cells, GlcNAc was still incorporated into O-glycans. However, in the case of HepG2 cells, no qualitative changes in N-glycans between WT and SLC35A3 knockout cells nor between SLC35A2 knockout and double-knockout cells were observed. These findings suggest that SLC35A3 may not be the primary UDP-GlcNAc transporter and/or different mechanisms of UDP-GlcNAc transport into the Golgi apparatus may exist.     

Pancreatic ductal adenocarcinomas from Mexican patients present a distinct genomic mutational pattern

Molecular Biology Reports - Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers in humans, with less than 5% 5-year survival rate. PDAC is characterized by a small number of...