Synthetic circuits reveal how mechanisms of gene regulatory networks constrain evolution

Phenotypic variation is the raw material of adaptive Darwinian evolution. The phenotypic variation found in organismal development is biased towards certain phenotypes, but the molecular mechanisms behind such biases are still poorly understood. Gene regulatory networks have been proposed as one cause of constrained phenotypic variation. However, most pertinent evidence is theoretical rather than experimental. Here, we study evolutionary biases in two synthetic gene regulatory circuits expressed in Escherichia coli that produce a gene expression stripe—a pivotal pattern in embryonic development. The two parental circuits produce the same phenotype, but create it through different regulatory mechanisms. We show that mutations cause distinct novel phenotypes in the two networks and use a combination of experimental measurements, mathematical modelling and DNA sequencing to understand why mutations bring forth only some but not other novel gene expression phenotypes. Our results reveal that the regulatory mechanisms of networks restrict the possible phenotypic variation upon mutation. Consequently, seemingly equivalent networks can indeed be distinct in how they constrain the outcome of further evolution.     

Reproductive traits of the “ocellated killifish” Floridichthys polyommus Hubbs, 1936 (Pisces: Cyprinodontidae) inhabiting estuary of the Champotón River (Campeche,Mexico)

This study explores the reproductive biology and the influence of environmental factors on the reproductive traits of Floridichthys polyommus inhabiting the estuary of the Champoton River. The analysis included population structure, gonadal developmental stages, reproductive period, size at first sexual maturity, fecundity, sex ratio, and somatic indexes. A spatio‐temporal pattern in the distribution of the seven size classes was detected. The species showed a positive allometric growth. Sex ratio was 1.21:1 (females:males). Size at first maturity was 42.23 mm (females) and 47.8 mm (males). A higher absolute and relative fecundity was detected in Puente Champotón estuary (PCH) with lower salinities than those detected in the Delta (DE). The adaptive significance of this trait could be in response to the heavy osmotic constraint imposed by extreme salinities in the DE. Floridichthys polyommus prefers PCH for spawning, and as a result a higher number of ripe individuals was observed in this site, characterized by the highest nutrient levels, which can boost productivity and food availability for fish. Hepatosomatic index was negatively correlated with gonadosomatic index, suggesting energy transfer from the liver towards gamete production. Floridichthys polyommus exhibits an opportunistic strategy (early maturity, small oocytes, small clutches) suggesting that energy is allocated towards reproduction to compensate for the unpredictable hydrological conditions imposed by the estuary (particularly during the hurricane season). Floridichthys polyommus shows a reproductive strategy exhibited by fish living in unstable systems. A seasonal divergence was observed in the reproductive traits of F. polyommus, while differences between study sites were for fecundity.     

Studies on the mode of action of hygromycin B, an inhibitor of translocation in eukaryotes.

Hygromycin B is an unusual aminoglycoside antibiotic active against both prokaryotic and eukaryotic cells. Hygromycin B at 0.38 mM concentration completely halts yeast cell growth in rich media, presumably by preventing protein synthesis by cytoplasmic ribosomes. Polypeptide synthesis in cell-free extracts from rabbit reticulocytes, wheat germ and yeast is strongly blocked by low concentrations of hygromycin B. The antibiotic inhibits peptide chain elongation by yeast polysomes by preventing elongation factor EF-2-dependent translocation, although it does not affect either the formation of the EF-2-GTP-ribosome complex or the EF-2- and ribosome-dependent GTP hydrolysis which takes place uncoupled from translocation. The inhibition of translocation by hygromycin B might result from the stabilization of peptidyl-tRNA bound to the ribosomal acceptor site, since the stability of [3H]Phe-tRNA-EF-1-poly(U)-ribosome and [3H]Phe-tRNA-poly(U)-ribosome complexes is increased in the presence of hygromycin B. The inhibition of polyphenylalanine synthesis by reticulocyte ribosomes and enzymic translocation of peptidyl-tRNA by yeast polysomes can be reversed by increasing concentrations of EF-2 suggesting a relationship between the binding sites of EF-2 and hygromycin B on the ribosome. Neither non-enzymic translocation, that takes place in the presence of high potassium concentrations, nor the peptide bondforming step are affected by hygromycin B.     

Integrating ecosystem engineering and food webs

Ecosystem engineering, the physical modification of the environment by organisms, is a common and often influential process whose significance to food web structure and dynamics is largely unknown. In the light of recent calls to expand food web studies to include non‐trophic interactions, we explore how we might best integrate ecosystem engineering and food webs. We provide rationales justifying their integration and present a provisional framework identifying how ecosystem engineering can affect the nodes and links of food webs and overall organization; how trophic interactions with the engineer can affect the engineering; and how feedbacks between engineering and trophic interactions can affect food web structure and dynamics. We use a simple integrative food chain model to illustrate how feedbacks between the engineer and the food web can alter 1) engineering effects on food web dynamics, and 2) food web responses to extrinsic environmental perturbations. We identify four general challenges to integration that we argue can readily be met, and call for studies that can achieve this integration and help pave the way to a more general understanding of interaction webs in nature. Synthesis All species are affected by their physical environment. Because ecosystem engineering species modify the physical environment and belong to food webs, such species are potentially one of the most important bridges between the trophic and non‐trophic. We examine how to integrate the so far, largely independent research areas of ecosystem engineering and food webs. We present a conceptual framework for understanding how engineering can affect food webs and vice versa, and how feedbacks between the two alter ecosystem dynamics. With appropriate empirical studies and models, integration is achievable, paving the way to a more general understanding of interaction webs in nature.     

Results of the Implementation of a Pilot Model for the Bidirectional Screening and Joint Management of Patients with Pulmonary Tuberculosis and Diabetes Mellitus in Mexico

Background

Recently, the World Health Organisation and the International Union Against Tuberculosis and Lung Disease published a Collaborative Framework for the Care and Control of Tuberculosis (TB) and Diabetes (DM) (CFTB/DM) proposing bidirectional screening and joint management.

Objective

To evaluate the feasibility and effectiveness of the CFTB/DM in Mexico. Design. Prospective observational cohort. Setting. 15 primary care units in 5 states in Mexico. Participants: Patients aged ≥20 years diagnosed with DM or pulmonary TB who sought care at participating clinics. Intervention: The WHO/Union CFTB/DM was adapted and implemented according to official Mexican guidelines. We recruited participants from July 2012 to April 2013 and followed up until March 2014. Bidirectional screening was performed. Patients diagnosed with TB and DM were invited to receive TB treatment under joint management. Main outcome measures. Diagnoses of TB among DM, of DM among TB, and treatment outcomes among patients with DM and TB.

Results

Of 783 DM patients, 11 (1.4%) were unaware of their TB. Of 361 TB patients, 16 (4.4%) were unaware of their DM. 95 TB/DM patients accepted to be treated under joint management, of whom 85 (89.5%) successfully completed treatment. Multiple linear regression analysis with change in HbA1c and random capillary glucose as dependent variables revealed significant decrease with time (regression coefficients (β)  = −0.660, (95% confidence interval (CI), −0.96 to −0.35); and β = −1.889 (95% CI, −2.77 to −1.01, respectively)) adjusting by sex, age and having been treated for a previous TB episode. Patients treated under joint management were more likely to experience treatment success than patients treated under routine DM and TB programs as compared to historical (adjusted OR (aOR), 2.8, 95%CI 1.28–6.13) and same period (aOR 2.37, 95% CI 1.13–4.96) comparison groups.

Conclusions

Joint management of TB and DM is feasible and appears to improve clinical outcomes.     

Analysis of chemometric models applied to Raman spectroscopy for monitoring key metabolites of cell culture

The Food and Drug Administration (FDA) initiative of Process Analytical Technology (PAT) encourages the monitoring of biopharmaceutical manufacturing processes by innovative solutions. Raman spectroscopy and the chemometric modeling tool partial least squares (PLS) have been applied to this aim for monitoring cell culture process variables. This study compares the chemometric modeling methods of Support Vector Machine radial (SVMr), Random Forests (RF), and Cubist to the commonly used linear PLS model for predicting cell culture components—glucose, lactate, and ammonia. This research is performed to assess whether the use of PLS as standard practice is justified for chemometric modeling of Raman spectroscopy and cell culture data. Model development data from five small-scale bioreactors (2 × 1 L and 3 × 5 L) using two Chinese hamster ovary (CHO) cell lines were used to predict against a manufacturing scale bioreactor (2,000 L). Analysis demonstrated that Cubist predictive models were better for average performance over PLS, SVMr, and RF for glucose, lactate, and ammonia. The root mean square error of prediction (RMSEP) of Cubist modeling was acceptable for the process concentration ranges of glucose (1.437 mM), lactate (2.0 mM), and ammonia (0.819 mM). Interpretation of variable importance (VI) results theorizes the potential advantages of Cubist modeling in avoiding interference of Raman spectral peaks. Predictors/Raman wavenumbers (cm⁻¹) of interest for individual variables are X1139–X1141 for glucose, X846–X849 for lactate, and X2941–X2943 for ammonia. These results demonstrate that other beneficial chemometric models are available for use in monitoring cell culture with Raman spectroscopy.     

Environmental Influences on Dall's Sheep Survival

Understanding how species respond to environmental conditions can assist with conservation strategies and harvest management, especially in arctic and boreal regions that are experiencing rapid climate change. Although climatic influences on species distributions have been studied, broad-scale effects of climate on survival are less well known. We examined the interactive effects of meteorological and remotely sensed environmental variables on survival of Dall's sheep (Ovis dalli dalli) lambs and adults by synthesizing radio-telemetry data across their range. We used data from 9 studies of adult sheep and 2 studies of lambs that were conducted between 1997 and 2012 at sites spanning the species' range in Alaska, USA, and northwestern Canada. We obtained environmental variables throughout the range of Dall's sheep, including the normalized difference vegetation index (NDVI) from optical remote sensing, freeze-thaw frequency (FTF) from passive microwave remote sensing, and gridded climate variables such as snow water equivalent, temperature, and precipitation. We used Cox proportional hazard regression to investigate the effects of environmental variables recorded during summer, winter, and the previous winter on annual survival rates of Dall's sheep lambs and adults. Summer NDVI was the most influential environmental factor affecting lamb survival, with improved lamb survival occurring in years with a high maximum NDVI. Also, lamb predation by coyotes (Canis latrans) and golden eagles (Aquila chrysaetos) decreased substantially with increasing NDVI. The previous winter FTF had the strongest effect on adult survival, with decreased survival occurring after winters with high FTF. In addition, these remotely sensed environmental factors interacted with meteorological factors to affect survival, such that effects of winter temperature depended on summer NDVI and winter FTF. Warm winters increased lamb survival only when preceded by summers with high NDVI, and warm winters increased adult survival only when winter FTF was low. Thus, potential benefits of climate warming may be counteracted if wintertime freeze-thaw events markedly increase. Correlations among environmental variables across sites were low, and regional climate cycles such as the Pacific Decadal Oscillation (PDO) had weak effects, indicating substantial local variability in climatic conditions experienced by Dall's sheep across their range. These findings can help managers anticipate how Dall's sheep populations will respond to changes in local environmental conditions. Our results also highlight the utility of multiple remotely sensed environmental conditions for ungulate management, especially passive microwave products that provide valuable information on winter icing events. © 2020 The Wildlife Society.     

Assessment of lake sturgeon (Acipenser fulvescens) recruitment in a regulated spawning tributary of Rainy Lake,Ontario

Continued study of the relationship between lake sturgeon (Acipenser fulvescens) recruitment and hydroelectric dams and operations, in a variety of river systems and habitat types is needed to improve the ability to predict and monitor impacts of the hydroelectric industry on this species. Herein, we present results of a juvenile lake sturgeon study aimed at addressing concerns over an inferred lack of recruitment resulting from spawning downstream of a hydroelectric generating station (HGS). Two years of sampling (2015 and 2016) were conducted in five sections of a 41 km long reach of the Seine River, Ontario, a lake sturgeon spawning tributary of Rainy Lake. Using an established gillnetting method, deepwater habitat was targeted to capture juvenile lake sturgeon to assess relative abundance, recruitment (cohort strength), and growth. Deepwater habitat, defined as water depths >6 m in this system, comprised only 2.1% of the wetted area in this study area. Within these habitats, a total of 331 lake sturgeon capture events were observed over the 2-years study period. The majority of the lake sturgeon catch (85%) was comprised of age-0 to age-5 individuals (both sampling years combined). Although inter-annual variation in cohort strength was apparent, each cohort between 2006 and 2016 was represented. The spatial distribution of cohorts varied among river reaches with younger individuals (age-0 and age-1) occupying reaches proximal to the Sturgeon Falls HGS, and larger, older individuals (age-2 to age-5) occupying reaches further downstream. The rarity of age-6+ individuals can likely be explained by ongoing downstream redistribution of juveniles over time, out of the Seine River and into Rainy Lake. Growth of juvenile lake sturgeon captured in the Seine River was above average relative to conspecifics from other rivers in the Hudson Bay drainage. Unfortunately, baseline data sets required to facilitate comparisons of contemporary (post-construction Sturgeon Falls HGS) versus historical (i.e. pre- Sturgeon Falls HGS) lake sturgeon recruitment, or to evaluate the influence of the Seine River Water Management Plan (2004) on lake sturgeon recruitment, are lacking. However, juvenile Lake Sturgeon are more abundant in this system than what had been surmised based on recent studies which implemented random sampling. Results indicate that juvenile lake sturgeon may reside in spawning tributaries for several years (age-0 to age-5) prior to seeking alternate habitats and highlights the value of targeted sampling (i.e. by depth) along the flow axis of rivers downstream of spawning areas when assessing lake sturgeon recruitment patterns.