Recovering phylogenetic signal from frog mating calls

Goicoechea, N., De La Riva, I. & Padial, J. M. (2010). Recovering phylogenetic signal from frog mating calls. —Zoologica Scripta, 39, 411–154. Few studies have tried to analyse the phylogenetic information contained in frog mating calls. While some of those studies suggest that sexual selection deletes any phylogenetic signal, others indicate that frog calls do retain phylogenetic informative characters. Discordant results can be the outcome of disparate rates of character evolution and evolutionary plasticity of call characters in different groups of frogs, but also the result of applying different coding methods. No study to date has compared the relative performance of different coding methods in detecting phylogenetic signal in calls, hampering thus potential consilience between previous results. In this study, we analyse the strength of phylogenetic signal in 10 mating call characters of 11 related species of frogs belonging to three genera of Andean and Amazonian frogs (Anura: Terrarana: Strabomantidae). We use six quantitative characters (number of notes per call, note length, call length, number of pulses per note, fundamental frequency and dominant frequency) and four qualitative ones (presence/absence of: pseudopulses, frequency modulation in notes, amplitude modulation in notes and amplitude modulation in pulses). We code quantitative characters using four different coding and scaling methods: (i) gap‐coding, (ii) fixed‐scale, (iii) step‐matrix gap‐weighting with between‐characters scaling, and (iv) step‐matrix gap‐weighting with between‐states scaling. All four coding methods indicate that frog calls contain phylogenetic information. These results suggest that divergent selection on frog mating calls may not always be strong enough to eliminate phylogenetic signal. However, coding methods strongly affect the amount of recoverable information. Step‐matrix gap‐weighting with between‐characters scaling and gap‐coding are suggested as the best methods available for coding quantitative characters of frog calls. Also, our results indicate that the arbitrariness in selecting character states and the method for scaling transitions weights, rather than the number of character states, is what potentially biases phylogenetic analyses with quantitative characters.     

Detecting positive selection within genomes: the problem of biased gene conversion

The identification of loci influenced by positive selection is a major goal of evolutionary genetics. A popular approach is to perform scans of alignments on a genome-wide scale in order to find regions evolving at accelerated rates on a particular branch of a phylogenetic tree. However, positive selection is not the only process that can lead to accelerated evolution. Notably, GC-biased gene conversion (gBGC) is a recombination-associated process that results in the biased fixation of G and C nucleotides. This process can potentially generate bursts of nucleotide substitutions within hotspots of meiotic recombination. Here, we analyse the results of a scan for positive selection on genes on branches across the primate phylogeny. We show that genes identified as targets of positive selection have a significant tendency to exhibit the genomic signature of gBGC. Using a maximum-likelihood framework, we estimate that more than 20 per cent of cases of significantly elevated non-synonymous to synonymous substitution rates ratio (d_N/d_S), particularly in shorter branches, could be due to gBGC. We demonstrate that in some cases, gBGC can lead to very high d_N/d_S (more than 2). Our results indicate that gBGC significantly affects the evolution of coding sequences in primates, often leading to patterns of evolution that can be mistaken for positive selection.     

Bioactive motifs of agouti signal protein

The switch between the synthesis of eu- and pheomelanins is modulated by the interaction of two paracrine signaling molecules, alpha-melanocyte stimulating hormone (MSH) and agouti signal protein (ASP), which interact with melanocytes via the MSH receptor (MC1R). Comparison of the primary sequence of ASP with the known MSH pharmacophore provides no suggestion about the putative bioactive domain(s) of ASP. To identify such bioactive motif(s), we synthesized 15-mer peptides that spanned the primary sequence of ASP and determined their effects on the melanogenic activities of murine melanocytes. Northern and Western blotting were used, together with chemical analysis of melanins and enzymatic assays, to identify three distinct bioactive regions of ASP that down-regulate eumelanogenesis. The decrease in eumelanin production was mediated by down-regulation of mRNA levels for tyrosinase and other melanogenic enzymes, as occurs in vivo, and these effects were comparable to those elicited by intact recombinant ASP. Shorter peptides in those motifs were synthesized and their effects on melanogenesis were further investigated. The amino acid arginine, which is present in the MSH peptide pharmacophore (HFRW), is also in the most active domain of ASP (KVARP). Our data suggest that lysines and an arginine (in motifs such as KxxxxKxxR or KxxRxxxxK) are important for the bioactivity of ASP. Identification of the specific ASP epitope that interacts with the MC1R has potential pharmacological applications in treating dysfunctions of skin pigmentation.     

Recovering microbial genomes from metagenomes in hypersaline environments: The Good,the Bad and the Ugly

Current metagenomic tools allow the recovery of microbial genomes directly from the environment. This can be accomplished by binning metagenomic contigs according to their coverage and tetranucleotide frequency, followed by an estimation of the bin quality. The public availability of bioinformatics tools, together with the decreasing cost of next generation sequencing, are democratizing this powerful approach that is spreading from specialized research groups to the general public. Using metagenomes from hypersaline environments, as well as mock metagenomes composed of Archaea and Bacteria frequently found in these systems, we have analyzed the advantages and difficulties of the binning process in these extreme environments to tackle microbial population diversity. These extreme systems harbor relatively low species diversity but high intraspecific diversity, which can compromise metagenome assembly and therefore the whole binning process. The main goal is to compare the output of the binning process with what is previously known from the analyzed samples, based on years of study using different approaches. Several scenarios have been analyzed in detail: (i) a good quality bin from a species highly abundant in the environment; (ii) an intermediate quality bin with incongruences that can be solved by further analyses and manual curation, and (iii) a low-quality bin to investigate the failure to recover a very abundant microbial genome as well as some possible solutions. The latter can be considered the “great metagenomics anomaly” and is mainly due to assembly problems derived from the microdiversity of naturally co-existing populations in nature.     

Long inverted repeats in eukaryotic genomes: recombinogenic motifs determine genomic plasticity

Inverted repeats are unstable motifs in a genome, having a causal relation to fragment rearrangements and recombination events. We have investigated long inverted repeats (LIR) of > 30 bp in length in eukaryotic genomes to assess their contribution to genome stability. An algorithm was first designed for searching for LIRs with < 2 kb internal spacers and >85% identity (degree of homology between repeat copies of a LIR). There are much fewer LIRs in yeast, fruitfly, pufferfish and chicken than in Caenorhabditis elegans, zebrafish, frog and human. However, the high LIR frequencies do not necessarily imply high genome instability because of variant internal spacers and stem lengths and identities. From the collection of identified LIRs, we selected recombinogenic LIRs that had a short internal spacer and a high copy identity and were prone to induce high instability. We found that a relatively high proportion (5-9.8%) of the LIRs in C. elegans, zebrafish and frog were recombinogenic LIRs. In contrast, the proportions in human and mouse LIRs were quite low (0.4-1.1%) basically accounting for long internal spacers. We suggest that C. elegans, zebrafish and frog genomes are unstable in terms of the LIR frequency and the proportion of recombinogenic LIRs. For the other genomes, LIRs most likely have a minor impact.     

Composition strand asymmetries in prokaryotic genomes: mutational bias and biased gene orientation

Most prokaryotic genomes display strand compositional asymmetries, but the reasons for these biases remain unclear. When the distribution of gene orientation is biased, as it often is, this may induce a bias in composition, as codon frequencies are not identical. We show here that this effect can be estimated and removed, and that the residual base skews are the highest at third base codon positions and lower at first and second positions. This strongly suggests that compositional asymmetries result from 1) a replication-related mutational bias that is filtered through selective pressure and/or from 2) an uneven distribution of gene orientation. In most cases, the mutational bias alters the codon usage and amino acid frequencies of the leading and the lagging strand. However, these features are not ubiquitous amongst prokaryotes, and the biological reasons for them remain to be found.     

Pollux: platform independent error correction of single and mixed genomes

Background

Second-generation sequencers generate millions of relatively short, but error-prone, reads. These errors make sequence assembly and other downstream projects more challenging. Correcting these errors improves the quality of assemblies and projects which benefit from error-free reads.

Results

We have developed a general-purpose error corrector that corrects errors introduced by Illumina, Ion Torrent, and Roche 454 sequencing technologies and can be applied to single- or mixed-genome data. In addition to correcting substitution errors, we locate and correct insertion, deletion, and homopolymer errors while remaining sensitive to low coverage areas of sequencing projects. Using published data sets, we correct 94% of Illumina MiSeq errors, 88% of Ion Torrent PGM errors, 85% of Roche 454 GS Junior errors. Introduced errors are 20 to 70 times more rare than successfully corrected errors. Furthermore, we show that the quality of assemblies improves when reads are corrected by our software.

Conclusions

Pollux is highly effective at correcting errors across platforms, and is consistently able to perform as well or better than currently available error correction software. Pollux provides general-purpose error correction and may be used in applications with or without assembly.     

The evolution of biased codon and amino acid usage in nematode genomes

Despite the degeneracy of the genetic code, whereby different codons encode the same amino acid, alternative codons and amino acids are utilized nonrandomly within and between genomes. Such biases in codon and amino acid usage have been demonstrated extensively in prokaryote genomes and likely reflect a balance between the action of mutation, selection, and genetic drift. Here, we quantify the effects of selection and mutation drift as causes of codon and amino acid-usage bias in a large collection of nematode partial genomes from 37 species spanning approximately 700 Myr of evolution, as inferred from expressed sequence tag (EST) measures of gene expression and from base composition variation. Average G + C content at silent sites among these taxa ranges from 10% to 63%, and EST counts range more than 100-fold, underlying marked differences between the identities of major codons and optimal codons for a given species as well as influencing patterns of amino acid abundance among taxa. Few species in our sample demonstrate a dominant role of selection in shaping intragenomic codon-usage biases, and these are principally free living rather than parasitic nematodes. This suggests that deviations in effective population size among species, with small effective sizes among parasites, are partly responsible for species differences in the extent to which selection shapes patterns of codon usage. Nevertheless, a consensus set of optimal codons emerges that is common to most taxa, indicating that, with some notable exceptions, selection for translational efficiency and accuracy favors similar sets of codons regardless of the major codon-usage trends defined by base compositional properties of individual nematode genomes.     

Distribution of short interstitial telomere motifs in two plant genomes: putative origin and function

Background  

Short interstitial telomere motifs (telo boxes) are short sequences identical to plant telomere repeat units. They are observed within the 5' region of several genes over-expressed in cycling cells. In synergy with various cis-acting elements, these motifs participate in the activation of expression. Here, we have analysed the distribution of telo boxes within Arabidopsis thaliana and Oryza sativa genomes and their association with genes involved in the biogenesis of the translational apparatus.     

Accurate recognition of cis-regulatory motifs with the correct lengths in prokaryotic genomes

We present a new computational method for solving a classical problem, the identification problem of cis-regulatory motifs in a given set of promoter sequences, based on one key new idea. Instead of scoring candidate motifs individually like in all the existing motif-finding programs, our method scores groups of candidate motifs with similar sequences, called motif closures, using a P-value, which has substantially improved the prediction reliability over the existing methods. Our new P-value scoring scheme is sequence length independent, hence allowing direct comparisons among predicted motifs with different lengths on the same footing. We have implemented this method as a Motif Recognition Computer (MREC) program, and have extensively tested MREC on both simulated and biological data from prokaryotic genomes. Our test results indicate that MREC can accurately pick out the actual motif with the correct length as the best scoring candidate for the vast majority of the cases in our test set. We compared our prediction results with two motif-finding programs Cosmo and MEME, and found that MREC outperforms both programs across all the test cases by a large margin. The MREC program is available at http://csbl.bmb.uga.edu/~bingqiang/MREC1/.     

AIMIE: a web-based environment for detection and interpretation of significant sequence motifs in prokaryotic genomes

Motivation: Genomes contain biologically significant informationthat extends beyond that encoded in genes. Some of this informationrelates to various short dispersed repeats distributed throughoutthe genome. The goal of this work was to combine tools for detectionof statistically significant dispersed repeats in DNA sequenceswith tools to aid development of hypotheses regarding theirpossible physiological functions in an easy-to-use web-basedenvironment. Results: Ab Initio Motif Identification Environment (AIMIE)was designed to facilitate investigations of dispersed sequencemotifs in prokaryotic genomes. We used AIMIE to analyze theEscherichia coli and Haemophilus influenzae genomes in orderto demonstrate the utility of the new environment. AIMIE detectedrepeated extragenic palindrome (REP) elements, CRISPR repeats,uptake signal sequences, intergenic dyad sequences and severalother over-represented sequence motifs. Distributional patternsof these motifs were analyzed using the tools included in AIMIE. Availability: AIMIE and the related software can be accessedat our web site http://www.cmbl.uga.edu/software.html. Contact: mrazek{at}uga.edu Associate Editor: Alex Bateman     

Identification of DNA motifs implicated in maintenance of bacterial core genomes by predictive modeling

Bacterial biodiversity at the species level, in terms of gene acquisition or loss, is so immense that it raises the question of how essential chromosomal regions are spared from uncontrolled rearrangements. Protection of the genome likely depends on specific DNA motifs that impose limits on the regions that undergo recombination. Although most such motifs remain unidentified, they are theoretically predictable based on their genomic distribution properties. We examined the distribution of the “crossover hotspot instigator,” or Chi, in Escherichia coli, and found that its exceptional distribution is restricted to the core genome common to three strains. We then formulated a set of criteria that were incorporated in a statistical model to search core genomes for motifs potentially involved in genome stability in other species. Our strategy led us to identify and biologically validate two distinct heptamers that possess Chi properties, one in Staphylococcus aureus, and the other in several streptococci. This strategy paves the way for wide-scale discovery of other important functional noncoding motifs that distinguish core genomes from the strain-variable regions.     

Comprehensive analysis of distinctive polyketide and nonribosomal peptide structural motifs encoded in microbial genomes

We developed a highly accurate method to predict polyketide (PK) and nonribosomal peptide (NRP) structures encoded in microbial genomes. PKs/NRPs are polymers of carbonyl/peptidyl chains synthesized by polyketide synthases (PKS) and nonribosomal peptide synthetases (NRPS). We analyzed domain sequences corresponding to specific substrates and physical interactions between PKSs/NRPSs in order to predict which substrates (carbonyl/peptidyl units) are selected and assembled into highly ordered chemical structures. The predicted PKs/NRPs were represented as the sequences of carbonyl/peptidyl units to extract the structural motifs efficiently. We applied our method to 4529 PKSs/NRPSs and found 619 PKs/NRPs. We also collected 1449 PKs/NRPs whose chemical structures have been determined experimentally. The structural sequences were compared using the Smith-Waterman algorithm, and clustered into 271 clusters. From the compound clusters, we extracted 33 structural motifs that are significantly related with their bioactivities. We used the structural motifs to infer functions of 13 novel PKs/NRPs clusters produced by Pseudomonas spp. and Burkholderia spp. and found a putative virulence factor. The integrative analysis of genomic and chemical information given here will provide a strategy to predict the chemical structures, the biosynthetic pathways, and the biological activities of PKs/NRPs, which is useful for the rational design of novel PKs/NRPs.