Relaxed Functional Constraints Research Articles

Comparative Analysis of Mitochondrial Genome in Fishes

Mitochondrial genome is circular double stranded DNA molecule in most of the eukaryotic organism which is ubiquitously present in all cells as it associated with production of energy. The number of complete mitochondrial genomes sequenced has been increasing day by day. Till date, the complete mitochondrial genome more than 2000 fish species can be retrieved from the NCBI GenBank. Like other vertebrates, fish mitochondrial genome contains 37 genes which include 22 tRNAs, 2 rRNAs (12S & 16S), 13 protein coding genes and one noncoding regions (Control region). The length of fish mitochondrial genome varies based on the size of noncoding region (CR). The complete mitochondrial genome sequence length varies from species to species. This comparison of genome sequences envisage the variations are mostly in the control region as it is highly evolved than other genes of mitochondria. Its mutation rate is very high. Therefore more numbers of indels and intergenic sequences are found. It may be due to the genetic drift, relaxed functional constraint and reduced selection pressure in the evolution of fish mitogenomes.

Hox genes control homocercal caudal fin development and evolution.

Ancient bony fishes had heterocercal tails, like modern sharks and sturgeons, with asymmetric caudal fins and a vertebral column extending into an elongated upper lobe. Teleost fishes, in contrast, developed a homocercal tail characterized by two separate equal-sized fin lobes and the body axis not extending into the caudal fin. A similar heterocercal-to-homocercal transition occurs during teleost ontogeny, although the underlying genetic and developmental mechanisms for either transition remain unresolved. Here, we investigated the role of hox13 genes in caudal fin formation as these genes control posterior identity in animals. Analysis of expression profiles of zebrafish hox13 paralogs and phenotypes of CRISPR/Cas9-induced mutants showed that double hoxb13a and hoxc13a mutants fail to form a caudal fin. Furthermore, single mutants display heterocercal-like morphologies not seen since Mesozoic fossil teleosteomorphs. Relaxation of functional constraints after the teleost genome duplication may have allowed hox13 duplicates to neo- or subfunctionalize, ultimately contributing to the evolution of a homocercal tail in teleost fishes.

The Origin and Evolution of Plastid Genome Downsizing in Southern Hemispheric Cypresses (Cupressaceae).

Plastome downsizing is rare in photosynthetic seed plants. However, a large-scale study of five cupressophyte families (conifers II) indicated that the plastomes of some Cupressaceous genera are notably reduced and compact. Here, we enriched taxon sampling in Cupressaceae by adding plastomes of ten previously unreported genera to determine the origin, evolution, and consequences of plastome reduction in this family. We discovered that plastome downsizing is specific to Callitroideae (a Southern Hemispheric subfamily). Their plastomes are the smallest, encode the fewest plastid genes, and contain the fewest GC-end codons among Cupressaceae. We show that repeated tRNA losses and shrinkage of intergenic spacers together contributed to the plastome downsizing in Callitroideae. Moreover, our absolute nucleotide substitution rate analyses suggest relaxed functional constraints in translation-related plastid genes (clpP, infA, rpl, and rps), but not in photosynthesis- or transcription-related ones, of Callitris (the most diverse genus in Callitroideae). We hypothesize that the small and low-GC plastomes of Callitroideae emerged ca. 112–75 million years ago as an adaptation to increased competition with angiosperms on the Gondwana supercontinent. Our findings highlight Callitroideae as another case of plastome downsizing in photosynthetic seed plant lineages.

Evolution of the gene of the lectin, Phalera flavescens agglutinin (PFA)

The larva of the Japanese buff-tip moth (Phalera flavescens) has a novel N-acetyllactosamine-binding lectin, termed Phalera flavescens agglutinin (PFA). It has been suggested that the PFA gene evolved from a lysozyme gene through the loss of lysozyme activity sites. In this study, we investigated whether the PFA gene is evolving through positive selection or by relaxation of functional constraint. For this purpose, we sequenced several PFA-orthologous genes from P. assimilis, P. takasagoensis, and P. bucephala, and compared synonymous and non-synonymous changes to examine the state of natural selection. Although a number of non-synonymous changes were observed on several branches of the gene tree, these effects may have been caused by relaxation of a functional constraint rather than a mechanism of positive selection. Therefore, it is reasonable to conclude that a lysozyme gene in the common ancestor of the Phalera (genus) species lost its lysozyme activity and accumulated mutations during the evolution of the species. Although the actual function of PFA in the moth species is as yet unknown, the gene probably obtained the N-acetyllactosamine-specific lectin activity, collaterally.

Evidence for relaxed selection of mitogenome in rapid-flow cyprinids

BackgroundHypoxia adaptation is developed in many fish species, which helped them to habitat most of water bodies. However, fishes living under high oxygen concentration may lose this feature. Rapid flows provide high level and stable dissolved oxygen, which facilitate organism’s oxygen supply and energy production. Previous studies showed that fish species from rapid-flow habitats exhibited lower hypoxia tolerance compared with fish from intermediate- and slow-flow habitats. Mitochondrial genomes code 13 key components in oxidative phosphorylation pathway; these genes may be under relaxed selection in rapid-flow species.ObjectivesThe primary objectives of this study is to investigate the evolutionary patterns of the 13 mitochondrial OXPHOS genes among nine cyprinids from different water bodies and to test the hypotheses that mitochondrial OXPHOS genes may experience relaxed selection in rapid-flow habitats.MethodsWe classified nine cyprinid fish species into three groups based on their habitats: rapid-flow, intermediate-flow and slow-flow. To detect relaxed selections, we investigated the 13 protein-coding genes with codon evolution programs RELAX; to estimate evolutionary rates among the cyprinids, free-ratio model in Codeml program was applied; Branch-site models were applied to detect positive selection sites. The polymorphisms of homologous sites were evaluated with PROVEAN program and projected to 3D structure prediction of the proteins using SWISS-MODEL.ResultsWe found that nine out of the 13 genes are under relaxed selection in rapid-flow species. Furthermore, dN, dS and dN/dS are relatively increased when compared with those of intermediate-flow species. More amino acid polymorphic sites are presented in rapid-flow species than in intermediate- and slow-flow species. Furthermore, rapid-flow species had more deleterious substitutions than other groups. 3D structure prediction of these proteins and projection of the polymorphic sites indicated that these sites were randomly distributed, suggesting relaxed functional constraints of these proteins in rapid-flow species.ConclusionOur results suggest that mitochondrial genes are under relaxed selection in rapid-flow cyprinids.

Architecture-Based Behavioral Adaptation with Generated Alternatives and Relaxed Constraints

Software systems are increasingly required to autonomously adapt their architectural structures and/or behaviors to runtime environmental changes. However, existing architecture-based self-adaptation approaches mostly focus on structural adaptations within a predefined space of architectural alternatives (e.g., switching between two alternative services) while merely considering quality constraints (e.g., reliability and performance). In this paper, we propose a new architecture-based self-adaptation approach, which performs behavioral adaptations with automatically generated alternatives and supports relaxed functional constraints from the perspective of business value. Specifically, we propose a technique to automatically generate behavioral alternatives of a software system from the currently-employed architectural behavioral specification. We employ business value to comprehensively evaluate the behavioral alternatives while capturing the trade-offs among relaxed functional and quality constraints. We also introduce a genetic algorithm-based planning technique to efficiently search for the optimal (sometimes a near-optimal) behavioral alternative that can provide the best business value. The experimental study on an online order processing benchmark has shown promising results that the proposed approach can improve adaptation flexibility and business value with acceptable performance overhead.

Coevolution of Siglec-11 and Siglec-16 via gene conversion in primates

BackgroundSiglecs-11 and -16 are members of the sialic acid recognizing Ig-like lectin family, and expressed in same cells. Siglec-11 functions as an inhibitory receptor, whereas Siglec-16 exhibits activating properties. In humans, SIGLEC11 and SIGLEC16 gene sequences are extremely similar in the region encoding the extracellular domain due to gene conversions. Human SIGLEC11 was converted by the nonfunctional SIGLEC16P allele, and the converted SIGLEC11 allele became fixed in humans, possibly because it provides novel neuroprotective functions in brain microglia. However, the detailed evolutionary history of SIGLEC11 and SIGLEC16 in other primates remains unclear.ResultsWe analyzed SIGLEC11 and SIGLEC16 gene sequences of multiple primate species, and examined glycan binding profiles of these Siglecs. The phylogenetic tree demonstrated that gene conversions between SIGLEC11 and SIGLEC16 occurred in the region including the exon encoding the sialic acid binding domain in every primate examined. Functional assays showed that glycan binding preference is similar between Siglec-11 and Siglec-16 in all analyzed hominid species. Taken together with the fact that Siglec-11 and Siglec-16 are expressed in the same cells, Siglec-11 and Siglec-16 are regarded as paired receptors that have maintained similar ligand binding preferences via gene conversions. Relaxed functional constraints were detected on the SIGLEC11 and SIGLEC16 exons that underwent gene conversions, possibly contributing to the evolutionary acceptance of repeated gene conversions. The frequency of nonfunctional SIGLEC16P alleles is much higher than that of SIGLEC16 alleles in every human population.ConclusionsOur findings indicate that Siglec-11 and Siglec-16 have been maintained as paired receptors by repeated gene conversions under relaxed functional constraints in the primate lineage. The high prevalence of the nonfunctional SIGLEC16P allele and the fixation of the converted SIGLEC11 imply that the loss of Siglec-16 and the gain of Siglec-11 in microglia might have been favored during the evolution of human lineage.

Contribution of both positive selection and relaxation of selective constraints to degeneration of flyability during geese domestication.

Flyability is the most discrepant trait between modern-day geese and their wild ancestors, and the degeneration of flyability is a key marker of the successful domestication of wild geese. In light of the relatively short history of domestic geese, intense artificial selection is thought to play an important role in the degeneration of flyability. However, the underlying mechanism behind this phenomenon has seldom been investigated. In this study, we applied a molecular evolutionary approach to the evaluation of partial breeds of domestic geese in order to look for genes involved in the selection pressure toward degeneration of flyability. The haplotype networks, pairwise fixation index (FST) values, and analysis of molecular variance results all clearly illustrated a population variance between Landes geese and partial Chinese domestic geese. We also detected signatures of positive artificial selection in the COX2 and COX3 genes, and related selection in the HBB gene. Our results support the independent origins of partial European domestic geese and Chinese domestic geese. In addition, both positive artificial selection and the relaxation of functional constraints appeared to play important roles in the degeneration of flyability in domestic geese.

A Pathway-Centered Analysis of Pig Domestication and Breeding in Eurasia.

Ascertaining the molecular and physiological basis of domestication and breeding is an active area of research. Due to the current wide distribution of its wild ancestor, the wild boar, the pig (Sus scrofa) is an excellent model to study these processes, which occurred independently in East Asia and Europe ca. 9000 yr ago. Analyzing genome variability patterns in terms of metabolic pathways is attractive since it considers the impact of interrelated functions of genes, in contrast to genome-wide scans that treat genes or genome windows in isolation. To that end, we studied 40 wild boars and 123 domestic pig genomes from Asia and Europe when metabolic pathway was the unit of analysis. We computed statistical significance for differentiation (Fst) and linkage disequilibrium (nSL) statistics at the pathway level. In terms of Fst, we found 21 and 12 pathways significantly differentiated at a q-value < 0.05 in Asia and Europe, respectively; five were shared across continents. In Asia, we found six significant pathways related to behavior, which involved essential neurotransmitters like dopamine and serotonin. Several significant pathways were interrelated and shared a variable percentage of genes. There were 12 genes present in >10 significant pathways (in terms of Fst), comprising genes involved in the transduction of a large number of signals, like phospholipase PCLB1, which is expressed in the brain, or ITPR3, which has an important role in taste transduction. In terms of nSL, significant pathways were mainly related to reproductive performance (ovarian steroidogenesis), a similarly important target trait during domestication and modern animal breeding. Different levels of recombination cannot explain these results, since we found no correlation between Fst and recombination rate. However, we did find an increased ratio of deleterious mutations in domestic vs. wild populations, suggesting a relaxed functional constraint associated with the domestication and breeding processes. Purifying selection was, nevertheless, stronger in significantly differentiated pathways than in random pathways, mainly in Europe. We conclude that pathway analysis facilitates the biological interpretation of genome-wide studies. Notably, in the case of pig, behavior played an important role, among other physiological and developmental processes.

Evolution of the RH gene family in vertebrates revealed by brown hagfish (Eptatretus atami) genome sequences

In vertebrates, there are four major genes in the RH (Rhesus) gene family, RH, RHAG, RHBG, and RHCG. These genes are thought to have been formed by the two rounds of whole-genome duplication (2R-WGD) in the common ancestor of all vertebrates. In our previous work, where we analyzed details of the gene duplications process of this gene family, three nucleotide sequences belonging to this family were identified in Far Eastern brook lamprey (Lethenteron reissneri), and the phylogenetic positions of the genes were determined. Lampreys, along with hagfishes, are cyclostomata (jawless fishes), which is a sister group of gnathostomata (jawed vertebrates). Although those results suggested that one gene was orthologous to the gnathostome RHCG genes, we did not identify clear orthologues for other genes. In this study, therefore, we identified three novel cDNA sequences that belong to the RH gene family using de novo transcriptome analysis of another cyclostome: the brown hagfish (Eptatretus atami). We also determined the nucleotide sequences for the RHBG and RHCG genes in a red stingray (Dasyatis akajei), which belongs to the cartilaginous fishes. The phylogenetic tree showed that two brown hagfish genes, which were probably duplicated in the cyclostome lineage, formed a cluster with the gnathostome RHAG genes, whereas another brown hagfish gene formed a cluster with the gnathostome RHCG genes. We estimated that the RH genes had a higher evolutionary rate than the RHAG, RHBG, and RHCG genes. Interestingly, in the RHBG genes, only the bird lineage showed a higher rate of nonsynonymous substitutions. It is likely that this higher rate was caused by a state of relaxed functional constraints rather than positive selection nor by pseudogenization.

Conservative and compensatory evolution in oxidative phosphorylation complexes of angiosperms with highly divergent rates of mitochondrial genome evolution.

Interactions between nuclear and mitochondrial gene products are critical for eukaryotic cell function. Nuclear genes encoding mitochondrial-targeted proteins (N-mt genes) experience elevated rates of evolution, which has often been interpreted as evidence of nuclear compensation in response to elevated mitochondrial mutation rates. However, N-mt genes may be under relaxed functional constraints, which could also explain observed increases in their evolutionary rate. To disentangle these hypotheses, we examined patterns of sequence and structural evolution in nuclear- and mitochondrial-encoded oxidative phosphorylation proteins from species in the angiosperm genus Silene with vastly different mitochondrial mutation rates. We found correlated increases in N-mt gene evolution in species with fast-evolving mitochondrial DNA. Structural modeling revealed an overrepresentation of N-mt substitutions at positions that directly contact mutated residues in mitochondrial-encoded proteins, despite overall patterns of conservative structural evolution. These findings support the hypothesis that selection for compensatory changes in response to mitochondrial mutations contributes to the elevated rate of evolution in N-mt genes. We discuss these results in light of theories implicating mitochondrial mutation rates and mitonuclear coevolution as drivers of speciation and suggest comparative and experimental approaches that could take advantage of heterogeneity in rates of mtDNA evolution across eukaryotes to evaluate such theories.

Sequence analysis of the Ras-MAPK pathway genes SOS1, EGFR & GRB2 in silver foxes (Vulpes vulpes): candidate genes for hereditary hyperplastic gingivitis.

Hereditary hyperplastic gingivitis (HHG) is an autosomal recessive disease that presents with progressive gingival proliferation in farmed silver foxes. Hereditary gingival fibromatosis (HGF) is an analogous condition in humans that is genetically heterogeneous with several known autosomal dominant loci. For one locus the causative mutation is in the Son of sevenless homologue 1 (SOS1) gene. For the remaining loci, the molecular mechanisms are unknown but Ras pathway involvement is suspected. Here we compare sequences for the SOS1 gene, and two adjacent genes in the Ras pathway, growth receptor bound protein 2 (GRB2) and epidermal growth factor receptor (EGFR), between HHG-affected and unaffected foxes. We conclude that the known HGF causative mutation does not cause HHG in foxes, nor do the coding regions or intron-exon boundaries of these three genes contain any candidate mutations for fox gum disease. Patterns of molecular evolution among foxes and other mammals reflect high conservation and strong functional constraints for SOS1 and GRB2 but reveal a lineage-specific pattern of variability in EGFR consistent with mutational rate differences, relaxed functional constraints, and possibly positive selection.

Relaxed functional constraints on triplicate α-globin gene in the bank vole suggest a different evolutionary history from other rodents

Gene duplication plays an important role in the origin of evolutionary novelties, but the mechanisms responsible for the retention and functional divergence of the duplicated copy are not fully understood. The α-globin genes provide an example of a gene family with different numbers of gene duplicates among rodents. Whereas Rattus and Peromyscus each have three adult α-globin genes (HBA-T1, HBA-T2 and HBA-T3), Mus has only two copies. High rates of amino acid evolution in the independently derived HBA-T3 genes of Peromyscus and Rattus have been attributed to positive selection. Using RACE PCR, reverse transcription-PCR (RT-PCR) and RNA-seq, we show that another rodent, the bank vole Clethrionomys glareolus, possesses three transcriptionally active α-globin genes. The bank vole HBA-T3 gene is distinguished from each HBA-T1 and HBA-T2 by 20 amino acids and is transcribed 23- and 4-fold lower than HBA-T1 and HBA-T2, respectively. Polypeptides corresponding to all three genes are detected by electrophoresis, demonstrating that the translated products of HBA-T3 are present in adult erythrocytes. Patterns of codon substitution and the presence of low-frequency null alleles suggest a postduplication relaxation of purifying selection on bank vole HBA-T3.

Progressive Genetic Architecture of Human Skin Pigmentation

Skin pigmentation is an important human phenotypic trait whose regulation, in spite of recent advances, has not been fully understood. Global variation in skin pigmentation is one of the most striking examples of environmental adaptation in humans. From genome-wide association studies (GWAS), almost four hundreds loci have been identified as candidate genes in model organisms. Nevertheless, only few of them have been found to be significantly associated with human skin pigmentation differences in African, Asian, and European populations. Further, the evolutionary history of different pigmentation genes is rather complex: some loci have been subjected to strong positive selection, while others evolved under the relaxation of functional constraints in low ultraviolet (UV) environment.In this manuscript, we report the current genetic architecture implicated in human skin pigmentation. Eventually, we conclude that this complex trait is strived to a bright future.

Cytonuclear Interactions and Relaxed Selection Accelerate Sequence Evolution in Organelle Ribosomes

Many mitochondrial and plastid protein complexes contain subunits that are encoded in different genomes. In animals, nuclear-encoded mitochondrial proteins often exhibit rapid sequence evolution, which has been hypothesized to result from selection for mutations that compensate for changes in interacting subunits encoded in mutation-prone animal mitochondrial DNA. To test this hypothesis, we analyzed nuclear genes encoding cytosolic and organelle ribosomal proteins in flowering plants. The model angiosperm genus Arabidopsis exhibits low organelle mutation rates, typical of most plants. Nevertheless, we found that (nuclear-encoded) subunits of organelle ribosomes in Arabidopsis have higher amino acid sequence polymorphism and divergence than their counterparts in cytosolic ribosomes, suggesting that organelle ribosomes experience relaxed functional constraint. However, the observed difference between organelle and cytosolic ribosomes was smaller than in animals and could be partially attributed to rapid evolution in N-terminal organelle-targeting peptides that are not involved in ribosome function. To test the role of organelle mutation more directly, we used transcriptomic data from an angiosperm genus (Silene) with highly variable rates of organelle genome evolution. We found that Silene species with unusually fast-evolving mitochondrial and plastid DNA exhibited increased amino acid sequence divergence in ribosomal proteins targeted to the organelles but not in those that function in cytosolic ribosomes. Overall, these findings support the hypothesis that rapid organelle genome evolution has selected for compensatory mutations in nuclear-encoded proteins. We conclude that coevolution between interacting subunits encoded in different genomic compartments within the eukaryotic cell is an important determinant of variation in rates of protein sequence evolution.

Global Patterns of Diversity and Selection in Human Tyrosinase Gene

Global variation in skin pigmentation is one of the most striking examples of environmental adaptation in humans. More than two hundred loci have been identified as candidate genes in model organisms and a few tens of these have been found to be significantly associated with human skin pigmentation in genome-wide association studies. However, the evolutionary history of different pigmentation genes is rather complex: some loci have been subjected to strong positive selection, while others evolved under the relaxation of functional constraints in low UV environment. Here we report the results of a global study of the human tyrosinase gene, which is one of the key enzymes in melanin production, to assess the role of its variation in the evolution of skin pigmentation differences among human populations. We observe a higher rate of non-synonymous polymorphisms in the European sample consistent with the relaxation of selective constraints. A similar pattern was previously observed in the MC1R gene and concurs with UV radiation-driven model of skin color evolution by which mutations leading to lower melanin levels and decreased photoprotection are subject to purifying selection at low latitudes while being tolerated or even favored at higher latitudes because they facilitate UV-dependent vitamin D production. Our coalescent date estimates suggest that the non-synonymous variants, which are frequent in Europe and North Africa, are recent and have emerged after the separation of East and West Eurasian populations.

Evolution of homospermidine synthase in the convolvulaceae: a story of gene duplication, gene loss, and periods of various selection pressures.

Homospermidine synthase (HSS), the first pathway-specific enzyme of pyrrolizidine alkaloid biosynthesis, is known to have its origin in the duplication of a gene encoding deoxyhypusine synthase. To study the processes that followed this gene duplication event and gave rise to HSS, we identified sequences encoding HSS and deoxyhypusine synthase from various species of the Convolvulaceae. We show that HSS evolved only once in this lineage. This duplication event was followed by several losses of a functional gene copy attributable to gene loss or pseudogenization. Statistical analyses of sequence data suggest that, in those lineages in which the gene copy was successfully recruited as HSS, the gene duplication event was followed by phases of various selection pressures, including purifying selection, relaxed functional constraints, and possibly positive Darwinian selection. Site-specific mutagenesis experiments have confirmed that the substitution of sites predicted to be under positive Darwinian selection is sufficient to convert a deoxyhypusine synthase into a HSS. In addition, analyses of transcript levels have shown that HSS and deoxyhypusine synthase have also diverged with respect to their regulation. The impact of protein-protein interaction on the evolution of HSS is discussed with respect to current models of enzyme evolution.

H2A.Z Nucleosome Positioning Has No Impact on Genetic Variation in Drosophila Genome

Nucleosome occupancy results in complex sequence variation rate heterogeneity by either increasing mutation rate or inhibiting DNA repair in yeast, fish, and human. H2A.Z nucleosome is extensively involved in gene transcription activation and regulation. To test whether H2A.Z nucleosome has the similar impact on sequence variability in the Drosophila genome, we profiled the H2A.Z nucleosome occupancy and sequence variation rate at gene ends and splicing sites. Consistent with previous studies, H2A.Z nucleosome positioning helps to demarcate the borders of exons. Nucleosome occupancy is anticorrelated with sequence divergence rate in the regions flanking transcription start sites and splicing sites. However, there is no rate heterogeneity between the linker DNA and H2A.Z nucleosomal DNA regardless of nucleosome occupancy, fuzziness, positioning in promoter, coding, and intergenic regions, young or old genes. But the rate at intergenic nucleosomes and the flanking linker regions is higher than that at the genic counterparts. Further analyses found that the high sequence divergence rate in the promoter regions that are usually nucleosome depleted regions may be likely resulted from the high mutation rate in the enriched tandem repeats. Interestingly, within nucleosomes spanning splicing sites, sequence variability of nucleosomal DNA significantly increases from the end within exons to the other end protruding into introns. The relaxed functional constraint in introns contributes to the high rate of nucleosomal DNA residing in introns while the strict functional constraint in exons maintains the low rate of nucleosomal DNA residing in exons. Taken together, H2A.Z nucleosome occupancy has no effect on sequence variability of Drosophila genome, which is likely determined by local sequence composition and the concomitant selection pressure.

Molecular evolution of the pigmentation gene melanocortin-1 receptor in rodents

Adaptive variation in the melanocortin 1-receptor gene (MC1R), a key locus in melanogenesis, has been identified in some species of rodents. However, in others, MC1R has no causative role in pigmentation phenotypes despite their coat color variation. In this study, we characterized the rates and patterns of MC1R nucleotide and amino acid sequence evolution and, particularly, selective pressures in the separated domains of the protein using a comparative analysis of 43 species representing three major lineages of rodents with variable coat colors. We found high amino acid variation (44% of sites) throughout the protein. Most substitutions were observed in extracellular and transmembrane domains; the intracellular segment was conserved across species. Pairwise non-synonymous substitutions did not vary significantly in different domains among the rodent lineages - i.e., variation was not associated with phylogenetic distance. Phylogeny-based likelihood analysis suggested that purifying selection has mostly shaped the evolutionary course of MC1R. However, a high proportion of sites (27%) were under relaxation of functional constraints (ω = 0.38), and four sites (3, 14, 26, and 251) clearly evolved under positive selection (ω ≅ 2.9). Thus, our data indicate a high proportion of sites evolving under relaxed evolutionary constraints, which might indicate the evolvability of the system in the generation of adaptive changes in specific taxa in rodent lineages.

Expression, divergence and evolution of the caleosin gene family in Brassica rapa

Caleosins (CLO) are oil body-associated proteins encoded by a small gene family. To investigate the expression, functional diversity and evolutionary modes of CLO genes, we isolated and integrally analyzed in silico a total of 11 CLO genes from Brassica rapa. According to phylogeny and sequence analyses, 11 BrCLO genes were classified into 3 groups, and each group shared highly conserved sequence features. Syntenic analysis revealed that all members of the BrCLO gene family distributed on 7 chromosomes were expanded mainly by segmental duplications. Evolutionary analysis showed that CLO proteins were controlled by purifying selection in B. rapa. Interestingly, functional divergence studies indicated that site-specific relaxed functional constraints were present between the different clusters of caleosins. Expression pattern suggested that 6 BrCLO genes were potentially associated with oil body formation. Our findings provide valuable clues for an investigation of the evolutionary history and cellular functions of the CLO gene family in plants.