Impacts of depression subcase and case on all-cause mortality in older people: The findings from the multi-centre community-based cohort study in China.

OBJECTIVES: It is unclear whether and to what extent depression subcases and cases in older age were associated with all-cause mortality. Little is known about gender differences in the associations. We assess these in older Chinese. METHODS: We examined a random sample of 6124 participants aged ≥60 years across five provinces in China. They were interviewed using a standard method of the GMS-AGECAT to diagnose depression subcase and case and record sociodemographic and disease risk factors at baseline, and to follow up their vital status. We employed Cox regression models to determine all-cause mortality in relation to depression subcases and cases, with adjustment for important variables, including social support and co-morbidities. RESULTS: Over the 10-year follow-up, 928 deaths occurred. Compared to those without depression at baseline, participants with depression subcase (n = 196) and case (n = 264) had increased risk of mortality; adjusted hazard ratios (HRs) were 1.46 (95% CI 1.07-2.00) and 1.45 (1.10-1.91). The adjusted HRs in men were 1.15 (0.72-1.81) and 1.85 (1.22-2.81), and in women 1.87 (1.22-2.87) and 1.22 (0.83-1.77) respectively. In participants aged ≥65 years, the adjusted HRs were 1.12 (0.68-1.84) and 1.99 (1.28-3.10) in men, and 2.06 (1.32-2.24) and 1.41 (0.94-2.10) in women. Increased HR in depression subcases was higher in women than man (ratio of HRs was 1.84, p = 0.034). CONCLUSIONS: Older people with depression subcase could have increased all-cause mortality to a similar extent to those with depression case. More attention should be paid to subcases of depression in women to tackle gender inequalities and improve survival.

Insights into the base-pairing preferences of 8-oxoguanosine on the ribosome.

Of the four bases, guanine is the most susceptible to oxidation, which results in the formation of 8-oxoguanine (8-oxoG). In protein-free DNA, 8-oxodG adopts the syn conformation more frequently than the anti one. In the syn conformation, 8-oxodG base pairs with dA. The equilibrium between the anti and syn conformations of the adduct are known to be altered by the enzyme recognizing 8-oxodG. We previously showed that 8-oxoG in mRNA severely disrupts tRNA selection, but the underlying mechanism for these effects was not addressed. Here, we use miscoding antibiotics and ribosome mutants to probe how 8-oxoG interacts with the tRNA anticodon in the decoding center. Addition of antibiotics and introduction of error-inducing mutations partially suppressed the effects of 8-oxoG. Under these conditions, rates and/or endpoints of peptide-bond formation for the cognate (8-oxoG•C) and near-cognate (8-oxoG•A) aminoacyl-tRNAs increased. In contrast, the antibiotics had little effect on other mismatches, suggesting that the lesion restricts the nucleotide from forming other interactions. Our findings suggest that 8-oxoG predominantly adopts the syn conformation in the A site. However, its ability to base pair with adenosine in this conformation is not sufficient to promote the necessary structural changes for tRNA selection to proceed.

Decoding on the ribosome depends on the structure of the mRNA phosphodiester backbone.

During translation, the ribosome plays an active role in ensuring that mRNA is decoded accurately and rapidly. Recently, biochemical studies have also implicated certain accessory factors in maintaining decoding accuracy. However, it is currently unclear whether the mRNA itself plays an active role in the process beyond its ability to base pair with the tRNA. Structural studies revealed that the mRNA kinks at the interface of the P and A sites. A magnesium ion appears to stabilize this structure through electrostatic interactions with the phosphodiester backbone of the mRNA. Here we examined the role of the kink structure on decoding using a well-defined in vitro translation system. Disruption of the kink structure through site-specific phosphorothioate modification resulted in an acute hyperaccurate phenotype. We measured rates of peptidyl transfer for near-cognate tRNAs that were severely diminished and in some instances were almost 100-fold slower than unmodified mRNAs. In contrast to peptidyl transfer, the modifications had little effect on GTP hydrolysis by elongation factor thermal unstable (EF-Tu), suggesting that only the proofreading phase of tRNA selection depends critically on the kink structure. Although the modifications appear to have no effect on typical cognate interactions, peptidyl transfer for a tRNA that uses atypical base pairing is compromised. These observations suggest that the kink structure is important for decoding in the absence of Watson-Crick or G-U wobble base pairing at the third position. Our findings provide evidence for a previously unappreciated role for the mRNA backbone in ensuring uniform decoding of the genetic code.

CYP2D6 Genotype-guided Metoprolol Therapy in Cardiac Surgery Patients: Rationale and Design of the Pharmacogenetic-guided Metoprolol Management for Postoperative Atrial Fibrillation in Cardiac Surgery (PREEMPTIVE) Pilot Study.

OBJECTIVES: The Preemptive Pharmacogenetic-guided Metoprolol Management for Atrial Fibrillation in Cardiac Surgery (PREEMPTIVE) pilot trial aims to use existing institutional resources to develop a process for integrating CYP2D6 pharmacogenetic test results into the patient electronic health record, to develop an evidence-based clinical decision support tool to facilitate CYP2D6 genotype-guided metoprolol administration in the cardiac surgery setting, and to determine the impact of implementing this CYP2D6 genotype-guided integrated approach on the incidence of postoperative atrial fibrillation (AF), provider, and cost outcomes. DESIGN: One-arm Bayesian adaptive design clinical trial. SETTING: Single center, university hospital. PARTICIPANTS: The authors will screen (including CYP2D6 genotype) up to 600 (264 ± 144 expected under the adaptive design) cardiac surgery patients, and enroll up to 200 (88 ± 48 expected) poor, intermediate, and ultrarapid CYP2D6 metabolizers over a period of 2 years at a tertiary academic center. INTERVENTIONS: All consented and enrolled patients will receive the intervention of CYP2D6 genotype-guided metoprolol management based on CYP2D6 phenotype classified as a poor, intermediate, extensive (normal), or ultrarapid metabolizer. MEASUREMENTS AND MAIN RESULTS: The primary outcome will be the incidence of postoperative AF. Secondary outcomes relating to rates of CYP2D6 genotype-guided prescription changes, costs, lengths of stay, and implementation metrics also will be investigated. CONCLUSIONS: The PREEMPTIVE pilot study is the first perioperative pilot trial to provide essential information for the design of a future, large-scale trial comparing CYP2D6 genotype-guided metoprolol management with a nontailored strategy in terms of managing AF. In addition, secondary outcomes regarding implementation, clinical benefit, safety, and cost-effectiveness in patients undergoing cardiac surgery will be examined.

ARGONAUTE 6 bridges transposable element mRNA-derived siRNAs to the establishment of DNA methylation.

Transposable elements (TEs) generate mutations and chromosomal instability when active. To repress TE activity, eukaryotic cells evolved mechanisms to both degrade TE mRNAs into small interfering RNAs (siRNAs) and modify TE chromatin to epigenetically inhibit transcription. Since the populations of small RNAs that participate in TE post-transcriptional regulation differ from those that establish RNA-directed DNA methylation (RdDM), the mechanism through which transcriptionally active TEs transition from post-transcriptional RNAi regulation to chromatin level control has remained unclear. We have identified the molecular mechanism of a plant pathway that functions to direct DNA methylation to transcriptionally active TEs. We demonstrated that 21-22 nucleotide (nt) siRNA degradation products from the RNAi of TE mRNAs are directly incorporated into the ARGONAUTE 6 (AGO6) protein and direct AGO6 to TE chromatin to guide its function in RdDM. We find that this pathway functions in reproductive precursor cells to primarily target long centromeric high-copy transcriptionally active TEs for RdDM prior to gametogenesis. This study provides a direct mechanism that bridges the gap between the post-transcriptional regulation of TEs and the establishment of TE epigenetic silencing.

Structured on-the-job training to improve retention of newborn resuscitation skills: a national cohort Helping Babies Breathe study in Tanzania.

BACKGROUND: Newborn resuscitation is a life-saving intervention for birth asphyxia, a leading cause of neonatal mortality. Improving provider newborn resuscitation skills is critical for delivering quality care, but the retention of these skills has been a challenge. Tanzania implemented a national newborn resuscitation using the Helping Babies Breathe (HBB) training program to help address this problem. Our objective was to evaluate the effectiveness of two training approaches to newborn resuscitation skills retention implemented across 16 regions of Tanzania. METHODS: An initial training approach implemented included verbal instructions for participating providers to replicate the training back at their service delivery site to others who were not trained. After a noted drop in skills, the program developed structured on-the-job training guidance and included this in the training. The approaches were implemented sequentially in 8 regions each with nurses/ midwives, other clinicians and medical attendants who had not received HBB training before. Newborn resuscitation skills were assessed immediately after training and 4-6 weeks after training using a validated objective structured clinical examination, and retention, measured through degree of skills drop, was compared between the two training approaches. RESULTS: Eight thousand, three hundred and ninety-one providers were trained and assessed: 3592 underwent the initial training approach and 4799 underwent the modified approach. Immediately post-training, average skills scores were similar between initial and modified training groups: 80.5 and 81.3%, respectively (p-value 0.07). Both groups experienced statistically significant drops in newborn resuscitation skills over time. However, the modified training approach was associated with significantly higher skills scores 4-6 weeks post training: 77.6% among the modified training approach versus 70.7% among the initial training approach (p-value < 0.0001). Medical attendant cadre showed the greatest skills retention. CONCLUSIONS: A modified training approach consisting of structured OJT, guidance and tools improved newborn resuscitation skills retention among health care providers. The study results give evidence for including on-site training as part of efforts to improve provider performance and strengthen quality of care.

The Potential Global Distribution and Voltinism of the Japanese Beetle (Coleoptera: Scarabaeidae) Under Current and Future Climates.

Japanese beetle, Popillia japonica (Newman), is a severe invasive insect pest of turf, landscapes, and horticultural crops. It has successfully colonized much of the United States and has recently established in mainland Europe. The distribution and voltinism of P. japonica will undoubtedly change as a consequence of climate change, posing additional challenges to the management of this species. To assess these challenges, a process-oriented bioclimatic niche model for P. japonica was developed to examine its potential global distribution under current (1981-2010) and projected climatic conditions (2040-2059) using one emission scenario (representative concentration pathway [RCP] 8.5) and two global climate models, ACCESS1-0 and CNRM-CM5. Under current climatic conditions, the bioclimatic niche model agreed well with all credible distribution data. Model projections indicate a strong possibility of further range expansion throughout mainland Europe under both current and future climates. In North America, projected increases in temperature would enable northward range expansion across Canada while simultaneously shifting southern range limits in the United States. In Europe, the suitable range for P. japonica would increase by 23% by midcentury, especially across portions of the United Kingdom, Ireland, and Scandinavia. Under the RCP 8.5 scenario, cumulative growing degree-days increased, thereby reducing the probability of biannual life cycles in northern latitudes where they can occur, including Hokkaido, Japan, northeastern portions of the United States, and southern Ontario, Canada. The results of this study highlight several regions of increasing and emerging risk from P. japonica that should be considered routinely in ongoing biosecurity and pest management surveys.

Food for thought: obstacles to menu labelling in restaurants and cafeterias.

Menu labelling is recommended as a policy intervention to reduce obesity and diet-related disease. The present commentary considers the many challenges the restaurant industry faces in providing nutrition information on its menus. Barriers include lack of nutrition expertise, time, cost, availability of nutrition information for exotic ingredients, ability to provide accurate nutrition information, libel risk, customer dissatisfaction, limited space on the menu, menu variations, loss of flexibility in changing the menu, staff training and resistance of employees to change current practice. Health promotion specialists and academics involved in fieldwork must help restaurateurs find solutions to these barriers for menu labelling interventions to be widely implemented and successful. Practical support for small independent restaurants such as free or subsidised nutrition analysis, nutrition training for staff and menu design may also be necessary to encourage voluntary participation.

Cost analysis of large-scale implementation of the 'Helping Babies Breathe' newborn resuscitation-training program in Tanzania.

BACKGROUND: Helping Babies Breathe (HBB) has become the gold standard globally for training birth-attendants in neonatal resuscitation in low-resource settings in efforts to reduce early newborn asphyxia and mortality. The purpose of this study was to do a first-ever activity-based cost-analysis of at-scale HBB program implementation and initial follow-up in a large region of Tanzania and evaluate costs of national scale-up as one component of a multi-method external evaluation of the implementation of HBB at scale in Tanzania. METHODS: We used activity-based costing to examine budget expense data during the two-month implementation and follow-up of HBB in one of the target regions. Activity-cost centers included administrative, initial training (including resuscitation equipment), and follow-up training expenses. Sensitivity analysis was utilized to project cost scenarios incurred to achieve countrywide expansion of the program across all mainland regions of Tanzania and to model costs of program maintenance over one and five years following initiation. RESULTS: Total costs for the Mbeya Region were $202,240, with the highest proportion due to initial training and equipment (45.2%), followed by central program administration (37.2%), and follow-up visits (17.6%). Within Mbeya, 49 training sessions were undertaken, involving the training of 1,341 health providers from 336 health facilities in eight districts. To similarly expand the HBB program across the 25 regions of mainland Tanzania, the total economic cost is projected to be around $4,000,000 (around $600 per facility). Following sensitivity analyses, the estimated total for all Tanzania initial rollout lies between $2,934,793 to $4,309,595. In order to maintain the program nationally under the current model, it is estimated it would cost $2,019,115 for a further one year and $5,640,794 for a further five years of ongoing program support. CONCLUSION: HBB implementation is a relatively low-cost intervention with potential for high impact on perinatal mortality in resource-poor settings. It is shown here that nationwide expansion of this program across the range of health provision levels and regions of Tanzania would be feasible. This study provides policymakers and investors with the relevant cost-estimation for national rollout of this potentially neonatal life-saving intervention.

Health Representations, Perceived Valence, and Concept Associations for Symbols as Food Cues: A Mixed-Methods Approach.

Researchers have experimented with a range of point-of-purchase (PoP) interventions in supermarkets, restaurants, and cafeterias. In general, these interventions have employed written materials. This research tested symbols to visually summarize information about the (un)healthiness of food. Study one explored health representations and valence associated with the image of a heart, a bathroom scale, and a running shoe using qualitative field interviews (N = 1200). Study two explored accessibility of a priori concept associations for two of those images, stratified by valence, in a computerized response latency task (N = 40). Study one indicted that the heart was best linked to its intended theme "heart health." Concerning valence, the heart was seen as both positive and negative whereas the scale was less likely to be viewed as positive relative to the running shoe. In study two, the heart was linked to five of the six a priori concepts and there was evidence that three of these were more accessible. Overall, the heart was better linked to positive poles than negative ones. A heart symbol may be useful to prompt heart healthy choices at the PoP. There was evidence that a scale may bias choice away from undesirable foods.

The initiation of epigenetic silencing of active transposable elements is triggered by RDR6 and 21-22 nucleotide small interfering RNAs.

Transposable elements (TEs) are mobile fragments of DNA that are repressed in both plant and animal genomes through the epigenetic inheritance of repressed chromatin and expression states. The epigenetic silencing of TEs in plants is mediated by a process of RNA-directed DNA methylation (RdDM). Two pathways of RdDM have been identified: RNA Polymerase IV (Pol IV)-RdDM, which has been shown to be responsible for the de novo initiation, corrective reestablishment, and epigenetic maintenance of TE and/or transgene silencing; and RNA-dependent RNA Polymerase6 (RDR6)-RdDM, which was recently identified as necessary for maintaining repression for a few TEs. We have further characterized RDR6-RdDM using a genome-wide search to identify TEs that generate RDR6-dependent small interfering RNAs. We have determined that TEs only produce RDR6-dependent small interfering RNAs when transcriptionally active, and we have experimentally identified two TE subfamilies as direct targets of RDR6-RdDM. We used these TEs to test the function of RDR6-RdDM in assays for the de novo initiation, corrective reestablishment, and maintenance of TE silencing. We found that RDR6-RdDM plays no role in maintaining TE silencing. Rather, we found that RDR6 and Pol IV are two independent entry points into RdDM and epigenetic silencing that perform distinct functions in the silencing of TEs: Pol IV-RdDM functions to maintain TE silencing and to initiate silencing in an RNA Polymerase II expression-independent manner, while RDR6-RdDM functions to recognize active Polymerase II-derived TE mRNA transcripts to both trigger and correctively reestablish TE methylation and epigenetic silencing.

Relationship between FEV1/FVC and age in children with asthma.

INTRODUCTION: Forced expiratory volume in the first second (FEV1)/forced vital capacity (FVC) normally decreases through childhood, increases briefly during early adolescence, and then declines throughout life. The physiology behind this temporary increase during early adolescence is not well understood. The objective of this study was to determine if this pattern occurs in children with asthma. DESIGN: Single-center, cross-sectional, retrospective analysis of pulmonary function tests obtained over a 5-year period in children 5-18 years of age with persistent asthma. RESULTS: A total of 1793 patients satisfied all inclusion and exclusion criteria. The mean age (±SD) was 10.4 ± 3.8 years. Forty-eight percent were female. Mean FEV1/FVC was 0.83 ± 0.09. FEV1/FVC was lower at 5 years of age than in healthy children, declined from age 5 to 11 by 5.7% compared to 7.3% in healthy girls, and 5.8% compared to 9.4% in healthy boys. FEV1/FVC increased in early adolescence, but at age 16, was 5.6% lower in male children compared to healthy children, and 5.4% lower in females. The ratio was lower in obese children at all ages but demonstrated the same curvilinear shape as healthy children. In absolute terms, FEV1 grew proportionately more than FVC during early adolescence, so the ratio of FEV1/FVC increased during that period. The curvilinear shape of the curve remained in postbronchodilator testing, though significantly blunted. CONCLUSIONS: FEV1/FVC is lower in children with persistent asthma than healthy children, but the "Shepherd's Hook" pattern is preserved. This was true in obese patients with asthma, although their FEV1/FVC ratios were lower throughout all stages of childhood and adolescence.

Verbal Behavior Analysis of Teaching Story Recall to Children with Autism: A Replication and Extension.

Children with autism spectrum disorder (ASD) may struggle with verbal behavior related to recall in various contexts. However, relatively little research has evaluated methods for improving recall among this population, and even fewer from a verbal behavior perspective. One socially important set of skills that relies upon a behavioral repertoire of recall is applied reading skills, such as reading comprehension and story recall. Valentino et al. (2015) designed an intervention package to teach children with ASD to recall short stories and conceptualized the behavior as an intraverbal chain. The present study replicated and extended that study with three school-aged children with ASD using a multiple baseline design across stories. For some participants and some stories, story recall was mastered under less intensive intervention conditions than in the previous study. When it was necessary to implement the full intervention package, the effects largely replicated previous research. Improvements in recall were correlated with increases in correct answers to comprehension questions. These data have important implications for clinicians and educators providing reading and recall interventions to children with ASD. Results also have theoretical implications for verbal behavior accounts of memory and recall, and suggest several possible avenues for future research. Supplementary Information: The online version contains supplementary material available at 10.1007/s40616-023-00183-2.

Introduction to the Collection: Climate Change, Insect Pests, and Beneficial Arthropods in Production Systems.

Climate change is expected to alter pressure from insect pests and the abundance and effectiveness of insect pollinators across diverse agriculture and forestry systems. In response to warming, insects are undergoing or are projected to undergo shifts in their geographic ranges, voltinism, abundance, and phenology. Drivers include direct effects on the focal insects and indirect effects mediated by their interactions with species at higher or lower trophic levels. These climate-driven effects are complex and variable, sometimes increasing pest pressure or reducing pollination and sometimes with opposite effects depending on climatic baseline conditions and the interplay of these drivers. This special collection includes several papers illustrative of these biological effects on pests and pollinators. In addition, in response to or anticipating climate change, producers are modifying production systems by introducing more or different crops into rotations or as cover crops or intercrops or changing crop varieties, with potentially substantial effects on associated insect communities, an aspect of climate change that is relatively understudied. This collection includes several papers illustrating these indirect production system-level effects. Together, biological and management-related effects on insects comprise the necessary scope for anticipating and responding to the effects of climate change on insects in agriculture and forest systems.

Patterns of adverse childhood experiences and depressive symptom trajectories in young adults: A longitudinal study of college students in China.

Background: Adverse childhood experiences (ACEs) tend to cluster together in daily life, and most studies focus on the level of depression at certain points, but the dynamic process of depression is often neglected. Thus, research is urgently needed to explore the relationship between ACEs pattern and trajectory of depressive symptom levels at multiple time points in order to provides early targeted interventions to those who are most at risk. Objective: We aimed to explore patterns of ACEs, including types and timing, associated with depression trajectories in college students. Methods: A school-based health survey was used to collect data as part of a longitudinal study in two medical college in Anhui province, China. Questionnaires were issued to 3,662 participants aged 17-22 and recorded details of ACEs (types and timing) and depression. Latent class analysis (LCA) was used to identify "patterns" of ACEs type and timing. Depressive symptom trajectories employed latent class growth analysis (LCGA). Multiple logistic regressions were employed to evaluate the relationships between ACEs patterns and depressive symptom trajectories. Results: We identified five ACEs patterns: "High neglect/emotional abuse/community violence," "High neglect/emotional abuse," "High neglect/family dysfunction," "High neglect," "Low ACEs." We traced three depression trajectories: "High depressive symptom" "Moderate depressive symptom," "Low depressive symptom." "High neglect/emotional abuse/community violence," "High neglect/emotional abuse" and "High neglect/family dysfunction" demonstrated a high risk for "High depressive symptom" and "Moderate depressive symptom." "High neglect" showed a high risk for "Moderate depressive symptom" but not for "High depressive symptom" (P < 0.05). Conclusions: The findings address the need for a comprehensive consideration of exposure to childhood adversity associated with the risk of depression in young adults through identifying more problematic ACEs patterns amongst exposed children.

Modeling Rangeland Grasshopper (Orthoptera: Acrididae) Population Density Using a Landscape-Level Predictive Mapping Approach.

Since the mid-19th century, grasshoppers have posed a substantial threat to North American rangelands as well as adjacent croplands and have the potential to cost the economy millions of dollars in annual damages. The United States Department of Agriculture (USDA) Animal and Plant Health Inspection Service (APHIS) have gone to great lengths to ensure that rangeland grasshopper populations remain below an economic impact threshold across the western United States. However, current grasshopper forecasting efforts by the USDA are based solely on the previous year's grasshopper density and do not take region-specific environmental factors (e.g., climate and topography) into account. To better understand the effects of climate and landscape heterogeneity on rangeland grasshopper populations, we assessed the relationship between grasshopper density survey data from across 56 sites between 2007 and 2017 for four counties in north central Wyoming with 72 biologically relevant geographic information system (GIS)-based environmental variables. A regression model was developed to predict mean adult grasshopper density from 2012 to 2016, which was then used to forecast grasshopper density in 2017. The best-fit predictive model selected using Akaike's Information Criterion (AICc) explained 34.5% of the variation in mean grasshopper density from 2012 to 2016. October precipitation and past mean grasshopper density from 2007 to 2011 were among the best predictors of mean grasshopper density in 2012-2016. Our results also suggest that rangelands in central Sheridan County, southwest Johnson County, and southeast Washakie County are more prone to grasshopper outbreaks. Most importantly, this study demonstrated that both biotic and abiotic environmental variables influence grasshopper density and should be considered in future forecasting efforts.

The Benefits of Yoga in the Classroom: A Mixed-Methods Approach to the Effects of Poses and Breathing and Relaxation Techniques.

BACKGROUND: Disadvantaged youth in the United States are disproportionately likely to be more sedentary and obese and experience more stress than their counterparts with higher socioeconomic status. Yoga and breathing and relaxation techniques have positive effects on stress levels, physical activity levels, and behavior of school-aged children. AIMS: Using social cognitive theory to examine behavioral, personal, and environmental factors, the purpose of this pilot study was to examine the multilevel influences of a yoga-based classroom intervention on urban youth. METHODS: Using a mixed methodological quasi-experimental design, this pilot study included the third grade students (n = 40) at one urban elementary school. A survey contained stress, yoga behavior, and aggression scales. In addition, individual student interviews, a teacher interview, and classroom observations were conducted. RESULTS: Paired and independent sample t-tests showed pre/post differences in yoga participation both in and out of school for the intervention participants (P < 0.01). Qualitative analysis revealed three main themes: (1) increased use and enjoyment of yoga techniques, (2) behavioral changes both in/out of school, and (3) impact on personal factors. CONCLUSIONS: Findings suggest that urban classrooms should include yoga and mindfulness training as it contributes to daily student PA and also can be stress relieving, fun, calming, and easy to perform outside of school.

Alkylative damage of mRNA leads to ribosome stalling and rescue by trans translation in bacteria.

Similar to DNA replication, translation of the genetic code by the ribosome is hypothesized to be exceptionally sensitive to small chemical changes to its template mRNA. Here we show that the addition of common alkylating agents to growing cultures of Escherichia coli leads to the accumulation of several adducts within RNA, including N(1)-methyladenosine (m1A). As expected, the introduction of m1A to model mRNAs was found to reduce the rate of peptide bond formation by three orders of magnitude in a well-defined in vitro system. These observations suggest that alkylative stress is likely to stall translation in vivo and necessitates the activation of ribosome-rescue pathways. Indeed, the addition of alkylation agents was found to robustly activate the transfer-messenger RNA system, even when transcription was inhibited. Our findings suggest that bacteria carefully monitor the chemical integrity of their mRNA and they evolved rescue pathways to cope with its effect on translation.

A short translational ramp determines the efficiency of protein synthesis.

Translation initiation is a major rate-limiting step for protein synthesis. However, recent studies strongly suggest that the efficiency of protein synthesis is additionally regulated by multiple factors that impact the elongation phase. To assess the influence of early elongation on protein synthesis, we employed a library of more than 250,000 reporters combined with in vitro and in vivo protein expression assays. Here we report that the identity of the amino acids encoded by codons 3 to 5 impact protein yield. This effect is independent of tRNA abundance, translation initiation efficiency, or overall mRNA structure. Single-molecule measurements of translation kinetics revealed pausing of the ribosome and aborted protein synthesis on codons 4 and 5 of distinct amino acid and nucleotide compositions. Finally, introduction of preferred sequence motifs only at specific codon positions improves protein synthesis efficiency for recombinant proteins. Collectively, our data underscore the critical role of early elongation events in translational control of gene expression.