Repeated, Selection-Driven Genome Reduction of Accessory Genes in Experimental Populations

Genome reduction has been observed in many bacterial lineages that have adapted to specialized environments. The extreme genome degradation seen for obligate pathogens and symbionts appears to be dominated by genetic drift. In contrast, for free-living organisms with reduced genomes, the dominant force is proposed to be direct selection for smaller, streamlined genomes. Most variation in gene content for these free-living species is of “accessory” genes, which are commonly gained as large chromosomal islands that are adaptive for specialized traits such as pathogenicity. It is generally unclear, however, whether the process of accessory gene loss is largely driven by drift or selection. Here we demonstrate that selection for gene loss, and not a shortened genome, per se, drove massive, rapid reduction of accessory genes. In just 1,500 generations of experimental evolution, 80% of populations of Methylobacterium extorquens AM1 experienced nearly parallel deletions removing up to 10% of the genome from a megaplasmid present in this strain. The absence of these deletion events in a mutation accumulation experiment suggested that selection, rather than drift, has dominated the process. Reconstructing these deletions confirmed that they were beneficial in their selective regimes, but led to decreased performance in alternative environments. These results indicate that selection can be crucial in eliminating unnecessary genes during the early stages of adaptation to a specialized environment

The Mechanism for RNA Recognition by ANTAR Regulators of Gene Expression

ANTAR proteins are widespread bacterial regulatory proteins that have RNA–binding output domains and utilize antitermination to control gene expression at the post-initiation level. An ANTAR protein, EutV, regulates the ethanolamine-utilization genes (eut) in Enterococcus faecalis. Using this system, we present genetic and biochemical evidence of a general mechanism of antitermination used by ANTARs, including details of the antiterminator structure. The novel antiterminator structure consists of two small hairpins with highly conserved terminal loop residues, both features being essential for successful antitermination. The ANTAR protein dimerizes and associates with its substrate RNA in response to signal-induced phosphorylation. Furthermore, bioinformatic searches using this conserved antiterminator motif identified many new ANTAR target RNAs in phylogenetically diverse bacterial species, some comprising complex regulons. Despite the unrelatedness of the species in which they are found, the majority of the ANTAR–associated genes are thematically related to nitrogen management. These data suggest that the central tenets for gene regulation by ANTAR antitermination occur widely in nature to specifically control nitrogen metabolism

Activation of EGFR/ERBB2 via Pathways Involving ERK1/2, P38 MAPK, AKT and FOXO Enhances Recovery of Diabetic Hearts from Ischemia-Reperfusion Injury

This study characterized the effects of diabetes and/or ischemia on epidermal growth factor receptor, EGFR, and/or erbB2 signaling pathways on cardiac function. Isolated heart perfusion model of global ischemia was used to study the effect of chronic inhibition or acute activation of EGFR/erbB2 signaling on cardiac function in a rat model of type-1 diabetes. Induction of diabetes with streptozotocin impaired recovery of cardiac function (cardiac contractility and hemodynamics) following 40 minutes of global ischemia in isolated hearts. Chronic treatment with AG825 or AG1478, selective inhibitors of erbB2 and EGFR respectively, did not affect hyperglycemia but led to an exacerbation whereas acute administration of the EGFR ligand, epidermal growth factor (EGF), led to an improvement in cardiac recovery in diabetic hearts. Diabetes led to attenuated dimerization and phosphorylation of cardiac erbB2 and EGFR receptors that was associated with reduced signaling via extracellular-signal-regulated kinase 1/2 (ERK1/2), p38 mitogen activated protein (MAP) kinase and AKT (protein kinase B). Ischemia was also associated with reduced cardiac signaling via these molecules whereas EGF-treatment opposed diabetes and/or ischemia induced changes in ERK1/2, p38 MAP kinase, and AKT-FOXO signaling. Losartan treatment improved cardiac function in diabetes but also impaired EGFR phosphorylation in diabetic heart. Co-administration of EGF rescued Losartan-mediated reduction in EGFR phosphorylation and significantly improved cardiac recovery more than with either agent alone. EGFR/erbB2 signaling is an important cardiac survival pathway whose activation, particularly in diabetes, ischemia or following treatment with drugs that inhibit this cascade, significantly improves cardiac function. These findings may have clinical relevance particularly in the treatment of diabetes-induced cardiac dysfunction

The Epidermal Growth Factor Receptor Is Involved in Angiotensin II But Not Aldosterone/Salt-Induced Cardiac Remodelling

Experimental and clinical studies have shown that aldosterone/mineralocorticoid receptor (MR) activation has deleterious effects in the cardiovascular system; however, the signalling pathways involved in the pathophysiological effects of aldosterone/MR in vivo are not fully understood. Several in vitro studies suggest that Epidermal Growth Factor Receptor (EGFR) plays a role in the cardiovascular effects of aldosterone. This hypothesis remains to be demonstrated in vivo. To investigate this question, we analyzed the molecular and functional consequences of aldosterone exposure in a transgenic mouse model with constitutive cardiomyocyte-specific overexpression of a mutant EGFR acting as a dominant negative protein (DN-EGFR). As previously reported, Angiotensin II-mediated cardiac remodelling was prevented in DN-EGFR mice. However, when chronic MR activation was induced by aldosterone-salt-uninephrectomy, cardiac hypertrophy was similar between control littermates and DN-EGFR. In the same way, mRNA expression of markers of cardiac remodelling such as ANF, BNF or β-Myosin Heavy Chain as well as Collagen 1a and 3a was similarly induced in DN-EGFR mice and their CT littermates. Our findings confirm the role of EGFR in AngII mediated cardiac hypertrophy, and highlight that EGFR is not involved in vivo in the damaging effects of aldosterone on cardiac function and remodelling

Reciprocal Sign Epistasis between Frequently Experimentally Evolved Adaptive Mutations Causes a Rugged Fitness Landscape

The fitness landscape captures the relationship between genotype and evolutionary fitness and is a pervasive metaphor used to describe the possible evolutionary trajectories of adaptation. However, little is known about the actual shape of fitness landscapes, including whether valleys of low fitness create local fitness optima, acting as barriers to adaptive change. Here we provide evidence of a rugged molecular fitness landscape arising during an evolution experiment in an asexual population of Saccharomyces cerevisiae. We identify the mutations that arose during the evolution using whole-genome sequencing and use competitive fitness assays to describe the mutations individually responsible for adaptation. In addition, we find that a fitness valley between two adaptive mutations in the genes MTH1 and HXT6/HXT7 is caused by reciprocal sign epistasis, where the fitness cost of the double mutant prohibits the two mutations from being selected in the same genetic background. The constraint enforced by reciprocal sign epistasis causes the mutations to remain mutually exclusive during the experiment, even though adaptive mutations in these two genes occur several times in independent lineages during the experiment. Our results show that epistasis plays a key role during adaptation and that inter-genic interactions can act as barriers between adaptive solutions. These results also provide a new interpretation on the classic Dobzhansky-Muller model of reproductive isolation and display some surprising parallels with mutations in genes often associated with tumors

Cardiovascular Response to Beta-Adrenergic Blockade or Activation in 23 Inbred Mouse Strains

We report the characterisation of 27 cardiovascular-related traits in 23 inbred mouse strains. Mice were phenotyped either in response to chronic administration of a single dose of the β-adrenergic receptor blocker atenolol or under a low and a high dose of the β-agonist isoproterenol and compared to baseline condition. The robustness of our data is supported by high trait heritabilities (typically H2>0.7) and significant correlations of trait values measured in baseline condition with independent multistrain datasets of the Mouse Phenome Database. We then focused on the drug-, dose-, and strain-specific responses to β-stimulation and β-blockade of a selection of traits including heart rate, systolic blood pressure, cardiac weight indices, ECG parameters and body weight. Because of the wealth of data accumulated, we applied integrative analyses such as comprehensive bi-clustering to investigate the structure of the response across the different phenotypes, strains and experimental conditions. Information extracted from these analyses is discussed in terms of novelty and biological implications. For example, we observe that traits related to ventricular weight in most strains respond only to the high dose of isoproterenol, while heart rate and atrial weight are already affected by the low dose. Finally, we observe little concordance between strain similarity based on the phenotypes and genotypic relatedness computed from genomic SNP profiles. This indicates that cardiovascular phenotypes are unlikely to segregate according to global phylogeny, but rather be governed by smaller, local differences in the genetic architecture of the various strains

A Tradeoff Drives the Evolution of Reduced Metal Resistance in Natural Populations of Yeast

Various types of genetic modification and selective forces have been implicated in the process of adaptation to novel or adverse environments. However, the underlying molecular mechanisms are not well understood in most natural populations. Here we report that a set of yeast strains collected from Evolution Canyon (EC), Israel, exhibit an extremely high tolerance to the heavy metal cadmium. We found that cadmium resistance is primarily caused by an enhanced function of a metal efflux pump, PCA1. Molecular analyses demonstrate that this enhancement can be largely attributed to mutations in the promoter sequence, while mutations in the coding region have a minor effect. Reconstruction experiments show that three single nucleotide substitutions in the PCA1 promoter quantitatively increase its activity and thus enhance the cells' cadmium resistance. Comparison among different yeast species shows that the critical nucleotides found in EC strains are conserved and functionally important for cadmium resistance in other species, suggesting that they represent an ancestral type. However, these nucleotides had diverged in most Saccharomyces cerevisiae populations, which gave cells growth advantages under conditions where cadmium is low or absent. Our results provide a rare example of a selective sweep in yeast populations driven by a tradeoff in metal resistance

Time-Resolved Transcriptome Analysis of Bacillus subtilis Responding to Valine, Glutamate, and Glutamine

Microorganisms can restructure their transcriptional output to adapt to environmental conditions by sensing endogenous metabolite pools. In this paper, an Agilent customized microarray representing 4,106 genes was used to study temporal transcript profiles of Bacillus subtilis in response to valine, glutamate and glutamine pulses over 24 h. A total of 673, 835, and 1135 amino-acid-regulated genes were identified having significantly changed expression at one or more time points in response to valine, glutamate, and glutamine, respectively, including genes involved in cell wall, cellular import, metabolism of amino-acids and nucleotides, transcriptional regulation, flagellar motility, chemotaxis, phage proteins, sporulation, and many genes of unknown function. Different amino acid treatments were compared in terms of both the global temporal profiles and the 5-minute quick regulations, and between-experiment differential genes were identified. The highlighted genes were analyzed based on diverse sources of gene functions using a variety of computational tools, including T-profiler analysis, and hierarchical clustering. The results revealed the common and distinct modes of action of these three amino acids, and should help to elucidate the specific signaling mechanism of each amino acid as an effector

Experimental Microbial Evolution of Extremophiles

Experimental microbial evolutions (EME) involves studying closely a microbial population after it has been through a large number of generations under controlled conditions (Kussell 2013). Adaptive laboratory evolution (ALE) selects for fitness under experimentally imposed conditions (Bennett and Hughes 2009; Dragosits and Mattanovich 2013). However, experimental evolution studies focusing on the contributions of genetic drift and natural mutation rates to evolution are conducted under non-selective conditions to avoid changes imposed by selection (Hindré et al. 2012). To understand the application of experimental evolutionary methods to extremophiles it is essential to consider the recent growth in this field over the last decade using model non-extremophilic microorganisms. This growth reflects both a greater appreciation of the power of experimental evolution for testing evolutionary hypotheses and, especially recently, the new power of genomic methods for analyzing changes in experimentally evolved lineages. Since many crucial processes are driven by microorganisms in nature, it is essential to understand and appreciate how microbial communities function, particularly with relevance to selection. However, many theories developed to understand microbial ecological patterns focus on the distribution and the structure of diversity within a microbial population comprised of single species (Prosser et al. 2007). Therefore an understanding of the concept of species is needed. A common definition of species using a genetic concept is a group of interbreeding individuals that is isolated from other such groups by barriers of recombination (Prosser et al. 2007). An alternative ecological species concept defines a species as set of individuals that can be considered identical in all relevant ecological traits (Cohan 2001). This is particularly important because of the abundance and deep phylogenetic complexity of microbial communities. Cohan postulated that “bacteria occupy discrete niches and that periodic selection will purge genetic variation within each niche without preventing divergence between the inhabitants of different niches”. The importance of gene exchange mechanisms likely in bacteria and archaea and therefore extremophiles, arises from the fact that their genomes are divided into two distinct parts, the core genome and the accessory genome (Cohan 2001). The core genome consists of genes that are crucial for the functioning of an organism and the accessory genome consists of genes that are capable of adapting to the changing ecosystem through gain and loss of function. Strains that belong to the same species can differ in the composition of accessory genes and therefore their capability to adapt to changing ecosystems (Cohan 2001; Tettelin et al. 2005; Gill et al. 2005). Additional ecological diversity exists in plasmids, transposons and pathogenicity islands as they can be easily shared in a favorable environment but still be absent in the same species found elsewhere (Wertz et al. 2003). This poses a major challenge for studying ALE and community microbial ecology indicating a continued need to develop a fitting theory that connects the fluid nature of microbial communities to their ecology (Wertz et al. 2003; Coleman et al. 2006). Understanding the nature and contribution of different processes that determine the frequencies of genes in any population is the biggest concern in population and evolutionary genetics (Prosser et al. 2007) and it is critical for an understanding of experimental evolution

Managing formalization to increase global team effectiveness and meaningfulness of work in multinational organizations

Global teams may help to integrate across locations, and yet, with formalized rules and procedures, responsiveness to those locations’ effectiveness, and the team members’ experiences of work as meaningful may suffer. We employ a mixed-methods approach to understand how the level and content of formalization can be managed to resolve these tensions in multinationals. In a sample of global teams from a large mining and resources organization operating across 44 countries, interviews, observations, and a quantitative 2-wave survey revealed a great deal of variability between teams in how formalization processes were enacted. Only those formalization processes that promoted knowledge sharing were instrumental in improving team effectiveness. Implementing rules and procedures in the set-up of the teams and projects, rather than during interactions, and utilizing protocols to help establish the global team as a source of identity increased this knowledge sharing. Finally, we found members’ personal need for structure moderated the effect of team formalization on how meaningful individuals found their work within the team. These findings have significant implications for theory and practice in multinational organizations