Three Microbial Musketeers of the Seas: Shewanella baltica,Aliivibrio fischeri and Vibrio harveyi,and Their Adaptation to Different Salinity Probed by a Proteomic Approach

Osmotic changes are common challenges for marine microorganisms. Bacteria have developed numerous ways of dealing with this stress, including reprogramming of global cellular processes. However, specific molecular adaptation mechanisms to osmotic stress have mainly been investigated in terrestrial model bacteria. In this work, we aimed to elucidate the basis of adjustment to prolonged salinity challenges at the proteome level in marine bacteria. The objects of our studies were three representatives of bacteria inhabiting various marine environments, Shewanella baltica, Vibrio harveyi and Aliivibrio fischeri. The proteomic studies were performed with bacteria cultivated in increased and decreased salinity, followed by proteolytic digestion of samples which were then subjected to liquid chromatography with tandem mass spectrometry analysis. We show that bacteria adjust at all levels of their biological processes, from DNA topology through gene expression regulation and proteasome assembly, to transport and cellular metabolism. The finding that many similar adaptation strategies were observed for both low- and high-salinity conditions is particularly striking. The results show that adaptation to salinity challenge involves the accumulation of DNA-binding proteins and increased polyamine uptake. We hypothesize that their function is to coat and protect the nucleoid to counteract adverse changes in DNA topology due to ionic shifts.     

Validation of Tuba1a as Appropriate Internal Control for Normalization of Gene Expression Analysis during Mouse Lung Development

The expression ratio between the analysed gene and an internal control gene is the most widely used normalization method for quantitative RT-PCR (qRT-PCR) expression analysis. The ideal reference gene for a specific experiment is the one whose expression is not affected by the different experimental conditions tested. In this study, we validate the applicability of five commonly used reference genes during different stages of mouse lung development. The stability of expression of five different reference genes (Tuba1a, Actb Gapdh, Rn18S and Hist4h4) was calculated within five experimental groups using the statistical algorithm of geNorm software. Overall, Tuba1a showed the least variability in expression among the different stages of lung development, while Hist4h4 and Rn18S showed the maximum variability in their expression. Expression analysis of two lung specific markers, surfactant protein C (SftpC) and Clara cell-specific 10 kDA protein (Scgb1a1), normalized to each of the five reference genes tested here, confirmed our results and showed that incorrect reference gene choice can lead to artefacts. Moreover, a combination of two internal controls for normalization of expression analysis during lung development will increase the accuracy and reliability of results.     

Binding Mode and Structure–Activity Relationships around Direct Inhibitors of the Nrf2–Keap1 Complex

An X‐ray crystal structure of Kelch‐like ECH‐associated protein (Keap1) co‐crystallised with (1S,2R)‐2‐[(1S)‐1‐[(1,3‐dioxo‐2,3‐dihydro‐1H‐isoindol‐2‐yl)methyl]‐1,2,3,4‐tetrahydroisoquinolin‐2‐carbonyl]cyclohexane‐1‐carboxylic acid (compound (S,R,S)‐ 1 a ) was obtained. This X‐ray crystal structure provides breakthrough experimental evidence for the true binding mode of the hit compound (S,R,S)‐ 1 a , as the ligand orientation was found to differ from that of the initial docking model, which was available at the start of the project. Crystallographic elucidation of this binding mode helped to focus and drive the drug design process more effectively and efficiently.     

Tributary confluences are dynamic thermal refuges for a juvenile salmonid in a warming river network

As rivers warm, cold‐water fish species may alleviate thermal stress by moving into localized thermal refuges such as cold‐water plumes created by cool tributary inflows. We quantified use of two tributary confluence plumes by juvenile steelhead, Oncorhynchus mykiss, throughout the summer, including how trout positioned themselves in relation to temperature within confluence plumes. At two confluences, Cedar and Elder creeks, along the South Fork Eel River, California, USA, we monitored temperatures using in situ logger grids throughout summer 2016. Fish were counted within confluences via snorkel surveys five times a day on 5 days at each site. We found diel and seasonal dependence on confluence use by steelhead, especially at the Cedar Creek confluence, where mainstem temperatures exceeded 28°C. At this site, fish moved into the confluence on the warmest days and warmest times of the day. Fish observed within the Cedar Creek confluence plume were most common in locations between 20–22°C, rather than the coldest locations (14.5°C). At Elder Creek, where mainstem temperatures remained below 24°C, there was little relationship between mainstem temperature and steelhead presence in the confluence plume. At both sites, steelhead distribution within plumes was influenced by spatial variation of temperature and mean temperature in surveyed grid cells. Our results show that cool tributaries flowing into warmer mainstem reaches (over 24°C) likely create important thermal refuges for juvenile steelhead. As mainstem rivers warm with climate change, cool‐water tributary inputs may become more important for sustaining cold‐water salmonids near the southern end of their range.     

Replication-Deficient Lymphocytic Choriomeningitis Virus-Vectored Vaccine Candidate for the Induction of T Cell Immunity against Mycobacterium tuberculosis

Mycobacterium tuberculosis (Mtb) represents a major burden to global health, and refined vaccines are needed. Replication-deficient lymphocytic choriomeningitis virus (rLCMV)-based vaccine vectors against cytomegalovirus have proven safe for human use and elicited robust T cell responses in a large proportion of vaccine recipients. Here, we developed an rLCMV vaccine expressing the Mtb antigens TB10.4 and Ag85B. In mice, rLCMV elicited high frequencies of polyfunctional Mtb-specific CD8 and CD4 T cell responses. CD8 but not CD4 T cells were efficiently boosted upon vector re-vaccination. High-frequency responses were also observed in neonatally vaccinated mice, and co-administration of rLCMV with Expanded Program of Immunization (EPI) vaccines did not result in substantial reciprocal interference. Importantly, rLCMV immunization significantly reduced the lung Mtb burden upon aerosol challenge, resulting in improved lung ventilation. Protection was associated with increased CD8 T cell recruitment but reduced CD4 T cell infiltration upon Mtb challenge. When combining rLCMV with BCG vaccination in a heterologous prime-boost regimen, responses to the rLCMV-encoded Mtb antigens were further augmented, but protection was not significantly different from rLCMV or BCG vaccination alone. This work suggests that rLCMV may show utility for neonatal and/or adult vaccination efforts against pulmonary tuberculosis.     

Shared book reading: When and how questions affect young children's word learning.

Shared book reading, and the conversation that accompanies it, can facilitate young children's vocabulary growth. To identify the features of extratextual questions that help 3-year-olds learn unfamiliar words during shared book reading, two experiments explored the impact of cognitive demand level, placement, and an approximation to scaffolding. Asking questions about target words improved children's comprehension and production of word–referent associations, and children with larger vocabularies learned more than children with smaller vocabularies. Neither the demand level nor placement of questions differentially affected word learning. However, an approximation to scaffolding, in which adults asked low demand questions when words first appeared and high demand questions later, did facilitate children's deeper understanding of word meanings as assessed with a definition task. These results are unique in experimentally demonstrating the value for word learning of shifting from less to more challenging input over time. Discussion focuses on why a scaffolding-like procedure improves children's acquisition of elaborated word meanings. (PsycINFO Database Record (c) 2010 APA, all rights reserved)