Evolutionary patterns in the antR-Cor gene in the dwarf dogwood complex (Cornus, Cornaceae)

The evolutionary pattern of the myc-like anthocyanin regulatory gene antR-Cor was examined in the dwarf dogwood species complex (Cornus Subgenus Arctocrania) that contains two diploid species (C. canadensis and C. suecica), their putative hybrids with intermediate phenotypes, and a tetraploid derivative (C. unalaschkensis). Full-length sequences of this gene (∼4 kb) were sequenced and characterized for 47 dwarf dogwood samples representing all taxa categories from 43 sites in the Pacific Northwest. Analysis of nucleotide diversity indicated departures from neutral evolution, due most likely to local population structure. Neighbor-joining and haplotype network analyses show that sequences from the tetraploid and diploid intermediates are much more strongly diverged from C. suecica than from C. canadensis, and that the intermediate phenotypes may represent an ancestral group to C. canadensis rather than interspecific hybrids. Seven amino acid mutations that are potentially linked to myc-like anthocyanin regulatory gene function correlate with petal colors differences that characterize the divergence between two diploid species and the tetraploid species in this complex. The evidence provides a working hypothesis for testing the role of the gene in speciation and its link to the petal coloration. Sequencing and analysis of additional nuclear genes will be necessary to resolve questions about the evolution of the dwarf dogwood complex.     

C1- and R-dependent expression of the maize Bz1 gene requires sequences with homology to mammalian myb and myc binding sites.

Tissue-specific expression of the maize anthocyanin Bronze-1 (Bz1) gene is controlled by the products of several regulatory genes. These include C1 or Pl and R or B that share homology to the myb proto-oncogenes and myc-like genes, respectively. Bz1 expression in embryo tissues is dependent on C1 and an R-sc allele of R. Transient expression from mutated and deleted versions of the Bz1 promoter fused to a luciferase reporter gene was measured in C1, Rscm2 embryos after gene transfer by microprojectiles. This analysis revealed that the sequences between -76 base pairs (bp) and -45 bp and a 9-bp AT-rich block between -88 bp and -80 bp were critical for Bz1 expression. The -76 bp to -45 bp region includes two short sequences that are homologous to the consensus binding sites of the myb- and myc-like proteins. Site-specific mutations of these "myb" and "myc" sequences reduced Bz1 expression to 10% and 1% of normal, respectively. Additionally, a trimer of a 38-bp oligonucleotide containing these myb and myc sites increased the expression of a cauliflower mosaic virus 35S minimal promoter by 26-fold. This enhancement was dependent on both C1 and R. Because the sites critical for Bz1 expression are homologous to the myb and myc consensus binding sequences and the C1 and R proteins share homology with the myb and myc products, respectively, we propose that C1 and R interact with the Bz1 promoter at these sites.     

Malate dehydrogenase: a useful phylogenetic marker for the genus Aeromonas

The reconstruction of correct genealogies among biological entities, the estimation of the divergence time between organisms or the study of the different events that occur along evolutionary lineages are not always based on suitable genes. For reliable results, it is necessary to look at full-length sequences of genes under stabilizing selection (neutral or purifying) and behaving as good molecular clocks. In bacteria it has been proved that the malate dehydrogenase gene (mdh) can be used to determine the inter- and intraspecies divergence, and hence this gene constitutes a potential marker for phylogeny and bacterial population genetics. We have sequenced the full-length mdh gene in 36 type and reference strains of Aeromonas. The species grouping obtained in the phylogenetic tree derived from mdh sequences was in agreement with that currently accepted for the genus Aeromonas. The maximum likelihood models applied to our sequences indicated that the mdh gene is highly conserved among the Aeromonas species and the main evolutionary force acting on it is purifying selection. Only two sites under potential diversifying selection were identified (T 108 and S 193). In order to determine if these two residues could have an influence on the MDH structure, we mapped them in a three-dimensional model constructed from the sequence of A. hydrophila using the human mitochondrial MDH as a template. The presence of purifying selection together with the linear relationship between substitutions and gene divergence makes the mdh an excellent candidate gene for a phylogeny of Aeromonas and probably for other bacterial groups.     

Molecular phylogenetic analysis ofChrysosplenium (Saxifragaceae) in Japan

The phylogeny of Japanese species ofChrysosplenium (Saxifragaceae) was examined using variation in DNA sequences. Sequences ofrbcL andmatK genes were compared for their feasibility for reconstructing the phylogeny ofChrysosplenium, and thematK sequences was found to give greater resolution. All but one of the 17 Japanese species have been examined formatK gene sequences and phylogenetic analysis of these data resulted in eight most parsimonious trees of 390 steps and a consistency index (Cl) of 0.823. The molecular phylogeny obtained was generally in agreement with Hara's (1957) classification based upon phenotypic similarity, although a conclusion needs extensive examination of the genus on a world-wide level. Using the phylogenetic data, character evolution was examined, especially in the characters traditionally used for grouping infrageneric taxa. Differentiation of opposite and alternate phyllotaxis appears to have occurred only once in the course of evolution ofChrysosplenium.     

Granule-bound starch synthase: structure, function, and phylogenetic utility

Interest in the use of low-copy nuclear genes for phylogenetic analyses of plants has grown rapidly, because highly repetitive genes such as those commonly used are limited in number. Furthermore, because low- copy genes are subject to different evolutionary processes than are plastid genes or highly repetitive nuclear markers, they provide a valuable source of independent phylogenetic evidence. The gene for granule-bound starch synthase (GBSSI or waxy) exists in a single copy in nearly all plants examined so far. Our study of GBSSI had three parts: (1) Amino acid sequences were compared across a broad taxonomic range, including grasses, four dicotyledons, and the microbial homologs of GBSSI. Inferred structural information was used to aid in the alignment of these very divergent sequences. The informed alignments highlight amino acids that are conserved across all sequences, and demonstrate that structural motifs can be highly conserved in spite of marked divergence in amino acid sequence. (2) Maximum-likelihood (ML) analyses were used to examine exon sequence evolution throughout grasses. Differences in probabilities among substitution types and marked among-site rate variation contributed to the observed pattern of variation. Of the parameters examined in our set of likelihood models, the inclusion of among-site rate variation following a gamma distribution caused the greatest improvement in likelihood score. (3) We performed cladistic parsimony analyses of GBSSI sequences throughout grasses, within tribes, and within genera to examine the phylogenetic utility of the gene. Introns provide useful information among very closely related species, but quickly become difficult to align among more divergent taxa. Exons are variable enough to provide extensive resolution within the family, but with low bootstrap support. The combined results of amino acid sequence comparisons, maximum-likelihood analyses, and phylogenetic studies underscore factors that might affect phylogenetic reconstruction. In this case, accommodation of the variable rate of evolution among sites might be the first step in maximizing the phylogenetic utility of GBSSI.      

Sequence diversity of a domesticated transposase gene, <Emphasis Type="Italic">MUG1</Emphasis>, in <Emphasis Type="Italic">Oryza</Emphasis> species

MUG1 is a MULE transposon-related domesticated gene in plants. We assessed the sequence diversity, neutrality, expression, and phylogenetics of the MUG1 gene among Oryza ssp. We found MUG1 expression in all tissues analyzed, with different levels in O. sativa. There were 408 variation sites in the 3886 bp of MUG1 locus. The nucleotide diversity of the MUG1 was higher than functionally known genes in rice. The nucleotide diversity (π) in the domains was lower than the average nucleotide diversity in whole coding region. The π values in nonsynonymous sites were lower than those of synonymous sites. Tajima D and Fu and Li D* values were mostly negative values, suggesting purifying selection in MUG1 sequences of Oryza ssp. Genome-specific variation and phylogenetic analyses show a general grouping of MUG1 sequences congruent with Oryza ssp. biogeography; however, our MUG1 phylogenetic results, in combination with separate B and D genome studies, might suggest an early divergence of the Oryza ssp. by continental drift of Gondwanaland. O. longistaminata MUG1 divergence from other AA diploids suggests that it might not be a direct ancestor of the African rice species. These authors contributed equally to this work.     

Cophylogeny and disparate rates of evolution in sympatric lineages of chewing lice on pocket gophers

Although molecular-based phylogenetic studies of hosts and parasites are increasingly common in the literature, no study to date has examined two congeneric lineages of parasites that live in sympatry on the same lineage of hosts. This study examines phylogenetic relationships among chewing lice (Phthiraptera: Trichodectidae) of the Geomydoecus coronadoi and Geomydoecus mexicanus species complexes and compares these to phylogenetic patterns in their hosts (pocket gophers of the rodent family Geomyidae). Sympatry of congeneric lice provides a natural experiment to test the hypothesis that closely related lineages of parasites will respond similarly to the same host. Sequence data from the mitochondrial COI and the nuclear EF-1alpha genes confirm that the two louse complexes are reciprocally monophyletic and that individual clades within each species complex parasitize a different species of pocket gopher. Phylogenetic comparisons reveal that both louse complexes show a significant pattern of cophylogeny with their hosts. Comparisons of rates of nucleotide substitution at 4-fold degenerate sites in the COI gene indicate that both groups of lice have significantly higher basal mutation rates than their hosts. The two groups of lice have similar basal rates of mutation, but lice of the G. coronadoi complex show significantly elevated rates of nucleotide substitution at all sites. These rate differences are hypothesized to result from population-level phenomena, such as effective population size, founder effects, and drift, that influence rates of nucleotide substitution.     

Molecular evolution and phylogenetic utility of Wolbachia ftsZ and wsp gene sequences with special reference to the origin of male-killing

A detailed assessment of the evolution and phylogenetic utility of two genes, ftsZ and wsp, was used to investigate the origin of male-killing Wolbachia, previously isolated from the ladybird Adalia bipunctata and the butterfly Acraea encedon. The analysis included almost all available sequences of B-group Wolbachia and two outgroup taxa and showed that (1) the two gene regions differ in phylogenetic utility, (2) sequence variation is here correlated with phylogenetic information content, (3) both genes show significant rate heterogeneity between lineages, (4) increased substitution rates are associated with homoplasy in the data, (5) wsp sequences of some taxa appear to be subject to positive selection, and (6) only a limited number of clades can be inferred with confidence due to either lack of phylogenetic information or the presence of homoplasy. With respect to the evolution of male-killing, the two genes nevertheless seemed to provide unbiased information. However, they consistently produce contradictory results. Current data therefore do not permit clarification of the origin of this behavior. In addition, A. bipunctata was found to be a host to two recently diverged strains of male-killing Wolbachia that showed increased substitution rates for both genes. Moreover, the wsp gene, which codes for an outer membrane protein, was found to be subject to positive selection in these taxa. These findings were postulated to be the product of high selection pressures due to antagonistic host-symbiont interactions in this ladybird species. In conclusion, our study demonstrates that the results of a detailed phylogenetic analysis, including characterization of the limitations of such an approach, can serve as a valuable basis for an understanding of the evolution of Wolbachia bacteria. Moreover, particular features of gene evolution, such as elevated substitution rates or the presence of positive selection, may provide information about the dynamics of Wolbachia-host associations.     

Comparative evolution of the mitochondrial cytochrome b gene and nuclear beta-fibrinogen intron 7 in woodpeckers

Most molecular phylogenetic studies of vertebrates have been based on DNA sequences of mitochondrial-encoded genes. MtDNA evolves rapidly and is thus particularly useful for resolving relationships among recently evolved groups. However, it has the disadvantage that all of the mitochondrial genes are inherited as a single linkage group so that only one independent gene tree can be inferred regardless of the number of genes sequenced. Introns of nuclear genes are attractive candidates for independent sources of rapidly evolving DNA: they are pervasive, most of their nucleotides appear to be unconstrained by selection, and PCR primers can be designed for sequences in adjacent exons where nucleotide sequences are conserved. We sequenced intron 7 of the beta-fibrinogen gene (beta-fibint7) for a diversity of woodpeckers and compared the phylogenetic signal and nucleotide substitution properties of this DNA sequence with that of mitochondrial-encoded cytochrome b (cyt b) from a previous study. A few indels (insertions and deletions) were found in the beta-fibint7 sequences, but alignment was not difficult, and the indels were phylogentically informative. The beta-fibint7 and cyt b gene trees were nearly identical to each other but differed in significant ways from the traditional woodpecker classification. Cyt b evolves 2.8 times as fast as beta-fibint7 (14. 0 times as fast at third codon positions). Despite its relatively slow substitution rate, the phylogenetic signal in beta-fibint7 is comparable to that in cyt b for woodpeckers, because beta-fibint7 has less base composition bias and more uniform nucleotide substitution probabilities. As a consequence, compared with cyt b, beta-fibint7 nucleotide sites are expected to enter more distinct character states over the course of evolution and have fewer multiple substitutions and lower levels of homoplasy. Moreover, in contrast to cyt b, in which nearly two thirds of nucleotide sites rarely vary among closely related taxa, virtually all beta-fibint7 nucleotide sites appear free of selective constraints, which increases informative sites per unit sequenced. However, the estimated gamma distribution used to model rate variation among sites suggests constraints on some beta-fibint7 sites. This study suggests that introns will be useful for phylogenetic studies of recently evolved groups.     

Molecular evolution of the hepatitis B virus genome

The hepatitis B virus (HBV) has a circular DNA genome of about 3,200 base pairs. Economical use of the genome with overlapping reading frames may have led to severe constraints on nucleotide substitutions along the genome and to highly variable rates of substitution among nucleotide sites. Nucleotide sequences from 13 complete HBV genomes were compared to examine such variability of substitution rates among sites and to examine the phylogenetic relationships among the HBV variants. The maximum likelihood method was employed to fit models of DNA sequence evolution that can account for the complexity of the pattern of nucleotide substitution. Comparison of the models suggests that the rates of substitution are different in different genes and codon positions; for example, the third codon position changes at a rate over ten times higher than the second position. Furthermore, substantial variation of substitution rates was detected even after the effects of genes and codon positions were corrected; that is, rates are different at different sites of the same gene or at the same codon position. Such rates after the correction were also found to be positively correlated at adjacent sites, which indicated the existence of conserved and variable domains in the proteins encoded by the viral genome. A multiparameter model validates the earlier finding that the variation in nucleotide conservation is not random around the HBV genome. The test for the existence of a molecular clock suggests that substitution rates are more or less constant among lineages. The phylogenetic relationships among the viral variants were examined. Although the data do not seem to contain sufficient information to resolve the details of the phylogeny, it appears quite certain that the serotypes of the viral variants do not reflect their genetic relatedness. Correspondence to: Z. Yang     

Enhancer Activity of Solitary Long Terminal Repeat of Human Endogenous Retrovirus K

The transient expression of the luciferase reporter gene was used to detect the tissue-specific enhancer activity of the solitary extraviral long terminal repeat (LTR) of the human endogenous retrovirus K (HERV-K). The LTR was previously mapped to the 19q13.2 locus. It contains a number of potential regulatory elements including TATA box, binding sites for some nuclear factors, and a polyadenylation signal. However, an analysis of the genomic sequences close to the LTR did not reveal any known genes or the expressed sequences (EST), whose functioning could be regulated by this LTR. The enhancer activity can be preserved in the solitary LTR due to its involvement in the long-range control of genome functioning or by the absence of functional disruptive mutations within the human-specific LTR, because it is of a relatively young evolutionary age.     

Analysis of among-site variation in substitution patterns

Substitution patterns among nucleotides are often assumed to be constant in phylogenetic analyses. Although variation in the average rate of substitution among sites is commonly accounted for, variation in the relative rates of specific types of substitution is not. Here, we review details of methodologies used for detecting and analyzing differences in substitution processes among predefined groups of sites. We describe how such analyses can be performed using existing phylogenetic tools, and discuss how new phylogenetic analysis tools we have recently developed can be used to provide more detailed and sensitive analyses, including study of the evolution of mutation and substitution processes. As an example we consider the mitochondrial genome, for which two types of transition deaminations (C⇒T and A⇒G) are strongly affected by single-strandedness during replication, resulting in a strand asymmetric mutation process. Since time spent single-stranded varies along the mitochondrial genome, their differential mutational response results in very different substitution patterns in different regions of the genome. Published: September 2, 2004.     

A rice <Emphasis Type="Italic">YABBY</Emphasis> gene, <Emphasis Type="Italic">OsYABBY4</Emphasis>, preferentially expresses in developing vascular tissue

Developmental gene families have diversified during land plant evolution. The primary role of YABBY gene family is promoting abaxial fate in model eudicot, Arabidopsis thaliana. However recent results suggest that roles of YABBY genes are not conserved in the angiosperms. In this paper, a rice YABBY gene was isolated, and its expression patterns were analyzed in detail. Sequence characterization and phylogenetic analyses showed the gene is OsYABBY4, which is group-classified into FIL/YAB3 subfamily. Beta-glucuronidase reporter assay and in situ analysis consistently revealed that OsYABBY4 was expressed in the meristems and developing vascular tissue of rice, predominantly in the phloem tissue, suggesting that the function of the rice gene is different from those of its counterparts in eudicots. OsYABBY4 may have been recruited to regulate the development of vasculature in rice. However, transgenic Arabidopsis plants ectopically expressing OsYABBY4 behaved very like those over-expressing FIL or YAB3 with abaxialized lateral organs, suggesting the OsYABBY4 protein domain is conserved with its Arabidopsis counterparts in sequences. Our results also indicate that the functional diversification of OsYABBY4 may be associated with the divergent spatial-temporal expression patterns, and YABBY family members may have preserved different expression regulatory systems and functions during the evolution of different kinds of species.     

Conserved noncoding sequences among cultivated cereal genomes identify candidate regulatory sequence elements and patterns of promoter evolution

Surveys for conserved noncoding sequences (CNS) among genes from monocot cereal species were conducted to assess the general properties of CNS in grass genomes and their correlation with known promoter regulatory elements. Initial comparisons of 11 orthologous maize-rice gene pairs found that previously defined regulatory motifs could be identified within short CNS but could not be distinguished reliably from random sequence matches. Among the different phylogenetic footprinting algorithms tested, the VISTA tool yielded the most informative alignments of noncoding sequence. VISTA was used to survey for CNS among all publicly available genomic sequences from maize, rice, wheat, barley, and sorghum, representing >300 gene comparisons. Comparisons of orthologous maize-rice and maize-sorghum gene pairs identified 20 bp as a minimal length criterion for a significant CNS among grass genes, with few such CNS found to be conserved across rice, maize, sorghum, and barley. The frequency and length of cereal CNS as well as nucleotide substitution rates within CNS were consistent with the known phylogenetic distances among the species compared. The implications of these findings for the evolution of cereal gene promoter sequences and the utility of using the nearly completed rice genome sequence to predict candidate regulatory elements in other cereal genes by phylogenetic footprinting are discussed.     

念珠藻属植物nifH基因的适应性进化分析

念珠藻(Nostoc)固氮过程关键在于固氮酶的催化,而固氮酶复合物中的铁蛋白(NifH)是由高度保守的nifH基因编码的,该基因是进化史上现存最古老的功能基因之一。该研究选取念珠藻属及近缘类群的nifH基因序列共40条,采用最大似然法构建系统发育树;运行PAML4.9软件,对nifH基因编码蛋白进行生物信息学分析,并使用分支模型、位点模型和分支-位点模型检测该基因的选择位点,探讨nifH基因的适应性进化特征。结果表明:(1)最大似然树显示内类群中该研究物种共分为6个分支(A、B、C、D、E和F),其中D和E是2个大的分支,每个大分支中又各包含2个特殊的小分支A、F和B、C,其中F分支包含新疆古尔班通古特沙漠采集到的9株念珠藻,A分支包含F分支及该研究测定序列的4株葛仙米,B分支包含本研究测定序列的4株地皮菜和3株未定种的念珠藻,C分支包含NCBI数据库中下载的5株念珠藻、鱼腥藻序列和本研究测定序列的1株念珠藻。(2)在所分析的3种进化模型中,仅通过分支-位点模型检测出14个统计学上显著的正选择位点,即1F、2S、3S、4T、5A、6F、7F、8I、9S、10C、17I、27Y、29D和31R位点,表明念珠藻属植物的nifH基因发生了适应性变化,分支-位点模型是研究藻类基因适应性进化较好的模型。