The Genetics of Adaptation for Eight Microvirid Bacteriophages

Theories of adaptive molecular evolution have recently experienced significant expansion, and their predictions and assumptions have begun to be subjected to rigorous empirical testing. However, these theories focus largely on predicting the first event in adaptive evolution, the fixation of a single beneficial mutation. To address long-term adaptation it is necessary to include new assumptions, but empirical data are needed for guidance. To empirically characterize the general properties of adaptive walks, eight recently isolated relatives of the single-stranded DNA (ssDNA) bacteriophage φX174 (family Microviridae) were adapted to identical selective conditions. Three of the eight genotypes were adapted in replicate, for a total of 11 adaptive walks. We measured fitness improvement and identified the genetic changes underlying the observed adaptation. Nearly all phages were evolvable; nine of the 11 lineages showed a significant increase in fitness. However, fitness plateaued quickly, and adaptation was achieved through only three substitutions on average. Parallel evolution was rampant, both across replicates of the same genotype as well as across different genotypes, yet adaptation of replicates never proceeded through the exact same set of mutations. Despite this, final fitnesses did not vary significantly among replicates. Final fitnesses did vary significantly across genotypes but not across phylogenetic groupings of genotypes. A positive correlation was found between the number of substitutions in an adaptive walk and the magnitude of fitness improvement, but no correlation was found between starting and ending fitness. These results provide an empirical framework for future adaptation theory

Uncoupling of Satellite DNA and Centromeric Function in the Genus Equus

In a previous study, we showed that centromere repositioning, that is the shift along the chromosome of the centromeric function without DNA sequence rearrangement, has occurred frequently during the evolution of the genus Equus. In this work, the analysis of the chromosomal distribution of satellite tandem repeats in Equus caballus, E. asinus, E. grevyi, and E. burchelli highlighted two atypical features: 1) several centromeres, including the previously described evolutionary new centromeres (ENCs), seem to be devoid of satellite DNA, and 2) satellite repeats are often present at non-centromeric termini, probably corresponding to relics of ancestral now inactive centromeres. Immuno-FISH experiments using satellite DNA and antibodies against the kinetochore protein CENP-A demonstrated that satellite-less primary constrictions are actually endowed with centromeric function. The phylogenetic reconstruction of centromere repositioning events demonstrates that the acquisition of satellite DNA occurs after the formation of the centromere during evolution and that centromeres can function over millions of years and many generations without detectable satellite DNA. The rapidly evolving Equus species gave us the opportunity to identify different intermediate steps along the full maturation of ENCs

Convergent evolution of chicken Z and human X chromosomes by expansion and gene acquisition

In birds, as in mammals, one pair of chromosomes differs between the sexes. In birds, males are ZZ and females ZW. In mammals, males are XY and females XX. Like the mammalian XY pair, the avian ZW pair is believed to have evolved from autosomes, with most change occurring in the chromosomes found in only one sex—the W and Y chromosomes1, 2, 3, 4, 5. By contrast, the sex chromosomes found in both sexes—the Z and X chromosomes—are assumed to have diverged little from their autosomal progenitors2. Here we report findings that challenge this assumption for both the chicken Z chromosome and the human X chromosome. The chicken Z chromosome, which we sequenced essentially to completion, is less gene-dense than chicken autosomes but contains a massive tandem array containing hundreds of duplicated genes expressed in testes. A comprehensive comparison of the chicken Z chromosome with the finished sequence of the human X chromosome demonstrates that each evolved independently from different portions of the ancestral genome. Despite this independence, the chicken Z and human X chromosomes share features that distinguish them from autosomes: the acquisition and amplification of testis-expressed genes, and a low gene density resulting from an expansion of intergenic regions. These features were not present on the autosomes from which the Z and X chromosomes originated but were instead acquired during the evolution of Z and X as sex chromosomes. We conclude that the avian Z and mammalian X chromosomes followed convergent evolutionary trajectories, despite their evolving with opposite (female versus male) systems of heterogamety. More broadly, in birds and mammals, sex chromosome evolution involved not only gene loss in sex-specific chromosomes, but also marked expansion and gene acquisition in sex chromosomes common to males and females.National Science Foundation (U.S.)Howard Hughes Medical Institut

Hidden heterochromatin: Characterization in the Rodentia species Cricetus cricetus, Peromyscus eremicus (Cricetidae) and Praomys tullbergi (Muridae)

The use of in situ restriction endonuclease (RE) (which cleaves DNA at specific sequences) digestion has proven to be a useful technique in improving the dissection of constitutive heterochromatin (CH), and in the understanding of the CH evolution in different genomes. In the present work we describe in detail the CH of the three Rodentia species, Cricetus cricetus, Peromyscus eremicus (family Cricetidae) and Praomys tullbergi (family Muridae) using a panel of seven REs followed by C-banding. Comparison of the amount, distribution and molecular nature of C-positive heterochromatin revealed molecular heterogeneity in the heterochromatin of the three species. The large number of subclasses of CH identified in Praomys tullbergi chromosomes indicated that the karyotype of this species is the more derived when compared with the other two genomes analyzed, probably originated by a great number of complex chromosomal rearrangements. The high level of sequence heterogeneity identified in the CH of the three genomes suggests the coexistence of different satellite DNA families, or variants of these families in these genomes

Analysis of genetic systems using experimental evolution and whole-genome sequencing

The application of whole-genome sequencing to the study of microbial evolution promises to reveal the complex functional networks of mutations that underlie adaptation. A recent study of parallel evolution in populations of Escherichia coli shows how adaptation involves both functional changes to specific proteins as well as global changes in regulation

Initial Mutations Direct Alternative Pathways of Protein Evolution

Whether evolution is erratic due to random historical details, or is repeatedly directed along similar paths by certain constraints, remains unclear. Epistasis (i.e. non-additive interaction between mutations that affect fitness) is a mechanism that can contribute to both scenarios. Epistasis can constrain the type and order of selected mutations, but it can also make adaptive trajectories contingent upon the first random substitution. This effect is particularly strong under sign epistasis, when the sign of the fitness effects of a mutation depends on its genetic background. In the current study, we examine how epistatic interactions between mutations determine alternative evolutionary pathways, using in vitro evolution of the antibiotic resistance enzyme TEM-1 β-lactamase. First, we describe the diversity of adaptive pathways among replicate lines during evolution for resistance to a novel antibiotic (cefotaxime). Consistent with the prediction of epistatic constraints, most lines increased resistance by acquiring three mutations in a fixed order. However, a few lines deviated from this pattern. Next, to test whether negative interactions between alternative initial substitutions drive this divergence, alleles containing initial substitutions from the deviating lines were evolved under identical conditions. Indeed, these alternative initial substitutions consistently led to lower adaptive peaks, involving more and other substitutions than those observed in the common pathway. We found that a combination of decreased enzymatic activity and lower folding cooperativity underlies negative sign epistasis in the clash between key mutations in the common and deviating lines (Gly238Ser and Arg164Ser, respectively). Our results demonstrate that epistasis contributes to contingency in protein evolution by amplifying the selective consequences of random mutations

The role of LINEs and CpG islands in dosage compensation on the chicken Z chromosome

Most avian Z genes are expressed more highly in ZZ males than ZW females, suggesting that chromosome-wide mechanisms of dosage compensation have not evolved. Nevertheless, a small percentage of Z genes are expressed at similar levels in males and females, an indication that a yet unidentified mechanism compensates for the sex difference in copy number. Primary DNA sequences are thought to have a role in determining chromosome gene inactivation status on the mammalian X chromosome. However, it is currently unknown whether primary DNA sequences also mediate chicken Z gene compensation status. Using a combination of chicken DNA sequences and Z gene compensation profiles of 310 genes, we explored the relationship between Z gene compensation status and primary DNA sequence features. Statistical analysis of different Z chromosomal features revealed that long interspersed nuclear elements (LINEs) and CpG islands are enriched on the Z chromosome compared with 329 other DNA features. Linear support vector machine (SVM) classifiers, using primary DNA sequences, correctly predict the Z compensation status for >60% of all Z-linked genes. CpG islands appear to be the most accurate classifier and alone can correctly predict compensation of 63% of Z genes. We also show that LINE CR1 elements are enriched 2.7-fold on the chicken Z chromosome compared with autosomes and that chicken chromosomal length is highly correlated with percentage LINE content. However, the position of LINE elements is not significantly associated with dosage compensation status of Z genes. We also find a trend for a higher proportion of CpG islands in the region of the Z chromosome with the fewest dosage-compensated genes compared with the region containing the greatest concentration of compensated genes. Comparison between chicken and platypus genomes shows that LINE elements are not enriched on sex chromosomes in platypus, indicating that LINE accumulation is not a feature of all sex chromosomes. Our results suggest that CpG islands are not randomly distributed on the Z chromosome and may influence Z gene dosage compensation status

Population-Based Resequencing of Experimentally Evolved Populations Reveals the Genetic Basis of Body Size Variation in Drosophila melanogaster

Body size is a classic quantitative trait with evolutionarily significant variation within many species. Locating the alleles responsible for this variation would help understand the maintenance of variation in body size in particular, as well as quantitative traits in general. However, successful genome-wide association of genotype and phenotype may require very large sample sizes if alleles have low population frequencies or modest effects. As a complementary approach, we propose that population-based resequencing of experimentally evolved populations allows for considerable power to map functional variation. Here, we use this technique to investigate the genetic basis of natural variation in body size in Drosophila melanogaster. Significant differentiation of hundreds of loci in replicate selection populations supports the hypothesis that the genetic basis of body size variation is very polygenic in D. melanogaster. Significantly differentiated variants are limited to single genes at some loci, allowing precise hypotheses to be formed regarding causal polymorphisms, while other significant regions are large and contain many genes. By using significantly associated polymorphisms as a priori candidates in follow-up studies, these data are expected to provide considerable power to determine the genetic basis of natural variation in body size

Phage Encoded H-NS: A Potential Achilles Heel in the Bacterial Defence System

The relationship between phage and their microbial hosts is difficult to elucidate in complex natural ecosystems. Engineered systems performing enhanced biological phosphorus removal (EBPR), offer stable, lower complexity communities for studying phage-host interactions. Here, metagenomic data from an EBPR reactor dominated by Candidatus Accumulibacter phosphatis (CAP), led to the recovery of three complete and six partial phage genomes. Heat-stable nucleoid structuring (H-NS) protein, a global transcriptional repressor in bacteria, was identified in one of the complete phage genomes (EPV1), and was most similar to a homolog in CAP. We infer that EPV1 is a CAP-specific phage and has the potential to repress up to 6% of host genes based on the presence of putative H-NS binding sites in the CAP genome. These genes include CRISPR associated proteins and a Type III restriction-modification system, which are key host defense mechanisms against phage infection. Further, EPV1 was the only member of the phage community found in an EBPR microbial metagenome collected seven months prior. We propose that EPV1 laterally acquired H-NS from CAP providing it with a means to reduce bacterial defenses, a selective advantage over other phage in the EBPR system. Phage encoded H-NS could constitute a previously unrecognized weapon in the phage-host arms race