Interspecific Comparison in the Frequency of Concerted Evolution at the Polyubiquitin Gene Locus

The polyubiquitin gene, encoding tandemly repeated multiple ubiquitins, constitutes a uniquitin gene subfamily. It has been demonstrated that polyubiquitin genes are subject to concerted evolution; namely, the individual ubiquitin coding units contained within a polyubiquitin gene are more similar to one another than they are to the ubiquitin coding units in the orthologous gene from other species. However there has been no comprehensive study on the concerted evolution of polyubiquitin genes in a wide range of species, because the relationships (orthologous or paralogous) among multiple polyubiquitin genes from different species have not been extensively analyzed yet. In this report, we present the results of analyzing the nucleotide sequence of polyubiquitin genes of mammals, available in the DDBJ/EMBL/GenBank nucleotide sequence databases, in which we found that there are two groups of polyubiquitin genes in an orthologous relationship. Based on this result, we analyzed the concerted evolution of the polyubiquitin gene in various species and compared the frequency of concerted evolutionary events interspecifically by taking into consideration that the rate of synonymous substitution at the polyubiquitin gene locus may vary depending on species. We found that the concerted evolutionary events in polyubiquitin genes have been more frequent in rats and Chinese hamsters than those in humans, cows, and sheep. The guinea pig polyubiquitin gene was an intermediate example. The frequency of concerted evolution in the mouse gene was unexpectedly low compared to that of other rodent genes. Received: 18 January 2000 / Accepted: 26 April 2000     

Rates of molecular evolution in nuclear genes of east Mediterranean scorpions

Scorpions of the genus Mesobuthus represent a useful terrestrial model system for studying molecular evolution. They are distributed on several Aegean islands and the adjacent mainland, they are believed to have low rates of dispersal, and evolutionary divergence dates of taxa are available based on biogeographic events that separated islands from each other and the mainland. Here, we present data on polymorphism and synonymous (Ks) and non-synonymous (Ka) substitution rates for nine nuclear protein-coding genes of two east Mediterranean scorpion species, Mesobuthus gibbosus and M. cyprius (Buthidae). Levels of polymorphism tend to be lower in populations from islands (mean nucleotide diversity pi = 0.0071 +/- 0.0028) than in mainland populations (mean pi = 0.0201 +/- 0.0085). By using linear regression of genetic divergence versus isolation time, we estimate Ks to be 3.17 +/- 1.54 per (site x 10(9) years), and Ka to be 0.39 +/- 0.94 per (site x 10(9) years). These estimates for both Ks and Ka are considerably lower than for many other invertebrates, such as Drosophila, and may be attributed to scorpions' mammal-like generation times (approximately 2 years) and low metabolic rates. Phylogenetic analysis using maximum likelihood revealed a phylogeny that is congruent with that expected based on biogeographic events and in which divergences at synonymous sites are proportional to the dates that the taxa are believed to have split. Tests of equality of branch lengths for the Cyprus and Crete lineages revealed that Ks-estimates are about the same in both lineages, as expected from the biogeographic events that separated the islands, but Ka was increased in the Cyprus lineage compared to the Cretan lineage.     

Evolutionarily conserved structure of the 3' non-translated region of a Chinese hamster polyubiquitin gene.

From a V79 Chinese hamster genomic library, we isolated a clone containing a polyubiquitin gene (designated as CHUB1), and determined its nucleotide sequence. The coding region of the CHUB1 gene consisted of five direct repeats of the ubiquitin unit with no spacer, followed by a single tyrosine residue. Northern hybridization analysis with a synthesized probe specific to the 3' non-translated region of the CHUB1 gene revealed that it codes for a 1.8 kb mRNA. An evident homology to the human polyubiquitin gene UbB and the chicken UbI gene was observed in the region corresponding to the full extent of the mature mRNA sequence, suggesting that these three genes belong to a common polyubiquitin gene subfamily, and that the sequence in the 3' non-translated region of the CHUB1 gene is unique to this subfamily.     

Ubiquitin genes as a paradigm of concerted evolution of tandem repeats

Summary Ubiquitin is remarkable for its ubiquitous distribution and its extreme protein sequence conservation. Ubiquitin genes comprise direct repeats of the ubiquitin coding unit with no spacers. The nucleotide sequences of several ubiquitin repeats from each of humans, chicken,Xenopus, Drosophila, barley, and yeast have recently been determined. By analysis of these data we show that ubiquitin is evolving more slowly than any other known protein, and that this (together with its gene organization) contributes to an ideal situation for the occurrence of concerted evolution of tandem repeats. By contrast, there is little evidence of between-cluster concerted evolution. We deduce that in ubiquitin genes, concerted evolution involves both unequal crossover and gene conversion, and that the average time since two repeated units within the polyubiquitin locus most recently shared a common ancestor is approximately 38 million years (Myr) in mammals, but perhaps only 11 Myr inDrosophila. The extreme conservatism of ubiquitin evolution also allows the inference that certain synonymous serine codons differing at the first two positions were probably mutated at single steps.     

The mouse polyubiquitin gene UbC is essential for fetal liver development, cell-cycle progression and stress tolerance

UbC is one of two stress-inducible polyubiquitin genes in mammals and is thought to supplement the constitutive UbA genes in maintaining cellular ubiquitin (Ub) levels during episodes of cellular stress. We have generated mice harboring a targeted disruption of the UbC gene. UbC(-/-) embryos die between embryonic days 12.5 and 14.5 in utero, most likely owing to a severe defect in liver cell proliferation. Mouse embryonic fibroblasts from UbC(-/-) embryos exhibit reduced growth rates, premature senescence, increased apoptosis and delayed cell-cycle progression, with slightly, but significantly, decreased steady-state Ub levels. UbC(-/-) fibroblasts are hypersensitive to proteasome inhibitors and heat shock, and unable to adequately increase Ub levels in response to these cellular stresses. Most, but not all of the UbC(-/-) phenotypes can be rescued by providing additional Ub from a poly hemagglutinin-tagged Ub minigene expressed from the Hprt locus. We propose that UbC is regulated by a process that senses Ub pool dynamics. These data establish that UbC constitutes an essential source of Ub during cell proliferation and stress that cannot be compensated by other Ub genes.     

Widespread positive selection in synonymous sites of mammalian genes

Evolution of protein sequences is largely governed by purifying selection, with a small fraction of proteins evolving under positive selection. The evolution at synonymous positions in protein-coding genes is not nearly as well understood, with the extent and types of selection remaining, largely, unclear. A statistical test to identify purifying and positive selection at synonymous sites in protein-coding genes was developed. The method compares the rate of evolution at synonymous sites (Ks) to that in intron sequences of the same gene after sampling the aligned intron sequences to mimic the statistical properties of coding sequences. We detected purifying selection at synonymous sites in approximately 28% of the 1,562 analyzed orthologous genes from mouse and rat, and positive selection in approximately 12% of the genes. Thus, the fraction of genes with readily detectable positive selection at synonymous sites is much greater than the fraction of genes with comparable positive selection at nonsynonymous sites, i.e., at the level of the protein sequence. Unlike other genes, the genes with positive selection at synonymous sites showed no correlation between Ks and the rate of evolution in nonsynonymous sites (Ka), indicating that evolution of synonymous sites under positive selection is decoupled from protein evolution. The genes with purifying selection at synonymous sites showed significant anticorrelation between Ks and expression level and breadth, indicating that highly expressed genes evolve slowly. The genes with positive selection at synonymous sites showed the opposite trend, i.e., highly expressed genes had, on average, higher Ks. For the genes with positive selection at synonymous sites, a significantly lower mRNA stability is predicted compared to the genes with negative selection. Thus, mRNA destabilization could be an important factor driving positive selection in nonsynonymous sites, probably, through regulation of expression at the level of mRNA degradation and, possibly, also translation rate. So, unexpectedly, we found that positive selection at synonymous sites of mammalian genes is substantially more common than positive selection at the level of protein sequences. Positive selection at synonymous sites might act through mRNA destabilization affecting mRNA levels and translation.     

Phosphoglucose isomerases of hagfish, zebrafish, gray mullet, toad, and snake, with reference to the evolution of the genes in vertebrates

Phosphoglucose isomerase (PGI) is a protein with multiple functions. To infer its structure changes and evolution in vertebrates, we cloned cDNAs encoding PGI genes from hagfish (Paramyxine yangi), gray mullet (Mugil cephalus), zebrafish (Danio rerio), toad (Bufo melanosticus), and snake (Boiga kraepelini). Only one PGI gene was cloned in each of hagfish, toad, and snake, but two PGI genes were found in zebrafish and gray mullet, respectively. The PGI of hagfish encodes 554 amino acids, in contrast to the PGIs of bonyfishes, toad, and snake which encode 553 amino acids and the PGIs of mammals which encode 558 amino acids. Among 558 aligned amino acid sites, there are 314 sites (56.27%) totally conserved. To see if diversifying selection acts on PGI amino acids of vertebrates, we calculated the pairwise ratio of nonsynonymous versus synonymous substitution per site (Ka/Ks) and the ratio of radical amino acid changes versus conservative amino acid changes per sites (dR/dC) between PGI sequences. The average pairwise ratio between nonsynonymous substitutions per nucleotide (Ka) and synonymous substitutions per nucleotide (Ks) among vertebrate PGI sequences equals 0.047 +/- 0.019. The average pairwise ratio between radical amino acid changes and conservative amino acid changes (dR/dC) among the vertebrate PGIs equal 0.938 +/- 0.158 for charge changes, 0.558 +/- 0.085 for polarity changes, and 0.465 +/- 0.0714 when both polarity and volume are considered. There is no amino acid within the vertebrate PGIs under diversifying selection as analyzed by the method of Yang et al. (2000b). The results suggest that the present vertebrate PGIs are at evolutionary stasis and are being subjected to intense purifying selection. The purifying selection is to maintain polarity and volume of the protein but not the charge groups of amino acids. Phylogenetic analysis reveals that vertebrate PGIs can be classified into three major groups: the mammalian, amphibian-reptilian, and teleostean PGIs. The gene tree suggests that the gene duplication event of PGI in bonyfishes occurred before diversification of Acanthopterygii but after the split of bonyfishes and tetrapods. The evolution of multiple functions of PGI is discussed.     

Structure and Evolution of Genes Encoding Polyubiquitin and Ubiquitin-like Proteins in Arabidopsis Thaliana Ecotype Columbia

The Arabidopsis thaliana ecotype Columbia ubiquitin gene family consists of 14 members that can be divided into three types of ubiquitin genes; polyubiquitin genes, ubiquitin-like genes and ubiquitin extension genes. The isolation and characterization of eight ubiquitin sequences, consisting of four polyubiquitin genes and four ubiquitin-like genes, are described here, and their relationships to each other and to previously identified Arabidopsis ubiquitin genes were analyzed. The polyubiquitin genes, UBQ3, UBQ10, UBQ11 and UBQ14, contain tandem repeats of the 228-bp ubiquitin coding region. Together with a previously described polyubiquitin gene, UBQ4, they differ in synonymous substitutions, number of ubiquitin coding regions, number and nature of nonubiquitin C-terminal amino acid(s) and chromosomal location, dividing into two subtypes; the UBQ3/UBQ4 and UBQ10/UBQ11/UBQ14 subtypes. Ubiquitin-like genes, UBQ7, UBQ8, UBQ9 and UBQ12, also contain tandem repeats of the ubiquitin coding region, but at least one repeat per gene encodes a protein with amino acid substitutions. Nucleotide comparisons, K(s) value determinations and neighbor-joining analyses were employed to determine intra- and intergenic relationships. In general, the rate of synonymous substitution is too high to discern related repeats. Specific exceptions provide insight into gene relationships. The observed nucleotide relationships are consistent with previously described models involving gene duplications followed by both unequal crossing-over and gene conversion events.     

广东人禽流感H5N1毒株M基因特性、进化和变异

通过对人禽流感H5N1毒株M基因序列的变异分析,揭示毒株M基因特征与进化。检测广东地区人禽流感H5N1毒株M基因核苷酸序列,同时检索全球人禽流感H5N1毒株M基因序列,采用DNAStar5.0软件对检索的人禽流感H5N1毒株M基因核苷酸序列进行比对和分析;并结合流行病学资料对变异毒株进行进化速度分析。结果发现,1997~2006年53株毒株M1基因和51株毒株M2核苷酸序列同源性均分成两组,1997年毒株为第一组(GⅠ),2003~2006年香港、越南、泰国、印尼、中国大陆、土耳其、伊拉克、阿塞拜疆、埃及毒株为第二组(GⅡ)。M1基因20个氨基酸位点置换,占7.94%(20/252),其中2003~2006年毒株M1基因有9个氨基酸位点不同于1997年毒株;M2基因22个氨基酸置换,占22.7%,其中2003~2006年毒株M2基因有4个氨基酸不同于1997年毒株。M2基因Ks值为26.8×10-6~42.6×10-6Nt/d,Ka值为4.39×10-6~6.98×10-6Nt/d;而M1基因的同义突变速度均远高于错义突变速度,显示M1基因受到机体免疫压力较小;检验发现M1基因进化存在负选择性压力。2003~2006年毒株M1基因通过氨基酸S224N置换,增加一个糖基化位点NSS224-226;而来自印尼的8株毒株M2基因发生C50F置换,引起蛋白二级结构改变。1997年中国香港人禽流感毒株自当时出现后,便未在以后人禽流感疫情中出现。2003~2006年毒株M1基因增加糖基化位点NSS224-226,可能与毒株致病性有关。人禽流感H5N1毒株M基因在自然界变异频繁,可能影响H5N1毒株的人-人传播能力。     

Phylogeny and substitution rates of angiosperm actin genes

Forty-four actin genes from five angiosperm species were PCR-amplified, cloned, and sequenced. Phylogenetic analysis of 34 of these actins, along with those previously published, indicates that angiosperm actin genes are monophyletic and underwent several duplications during evolution. Orthologues have been identified between Solanaceae species, as well as between Solanaceae species and soybean. These sequences were used to calculate nucleotide substitution rates. The synonymous rate (6.96 x 10(-9) substitutions/site/year) is similar to that of other nuclear protein-coding genes, but the nonsynonymous rate (0.19 x 10(-9) substitutions/site/year) is 6-19 times higher than that of mammalian actin genes. Relative rate tests indicate that actin genes are evolving at similar rates in monocots and in dicots. Evidence is also presented that some members of the maize actin multigene family have been involved in gene conversion events, that the potato genome contains 24 +/- 12 actin genes, and that potato and tomato diverged 11.6 +/- 3.6 MYA.      

Evolution of nucleotide substitutions and gene regulation in the amylase multigenes in Drosophila kikkawai and its sibling species

In order to determine evolutionary changes in gene regulation and the nucleotide substitution pattern in a multigene family, the amylase multigenes were characterized in Drosophila kikkawai and its sibling species. The nucleotide substitution pattern was investigated. Drosophila kikkawai has four amylase genes. The Amy1 and Amy2 genes are a head-to-head duplication in the middle of the B arm of the second chromosome, while the Amy3 and Amy4 genes are a tail-to-tail duplication near the centromere of the same chromosome. In the sibling species of D. kikkawai (Drosophila bocki, Drosophila leontia, and Drosophila lini), sequencing of the Amy1, Amy2, Amy3, and Amy4 genes revealed that the Amy1 and Amy2 gene group diverged from Amy3 and Amy4 after duplication. In the Amy1 and Amy2 genes, the divergent evolution occurred in the flanking regions; in contrast, the coding regions have evolved in concerted fashion. The electrophoretic pattern of AMY isozymes was also examined. In D. kikkawai and its siblings, two or three electrophoretically different isozymes are encoded by the Amy1 and Amy2 genes (S isozyme) and by the Amy3 and Amy4 genes (F (M) isozymes). The S and F (M) isozymes show different patterns of band intensity when larvae and flies were fed in different media. Amy1 and Amy2, which encode the S isozyme, are more strikingly regulated than Amy3 and Amy4, which encode the F (M) isozyme. The GC content and codon usage bias were higher for the Amy1 and Amy2 genes than for the Amy3 and Amy4 genes. Although the ratio of synonymous and replacement substitutions within the Amy1 and Amy2 gene group was not significantly different from that within the Amy3 and Amy4 gene group, the synonymous substitution rate in the lineage of Amy1 and Amy2 was lower than that of Amy3 and Amy4. In conclusion, after the first duplication but before speciation of four species, the synonymous substitution rate between the two lineages and the electrophoretic pattern of the isozymes encoded by them changed, although we do not know whether there was any evolutionary relationship between the two.     

Calibration of multiple poliovirus molecular clocks covering an extended evolutionary range

We have calibrated five different molecular clocks for circulating poliovirus based upon the rates of fixation of total substitutions (K(t)), synonymous substitutions (K(s)), synonymous transitions (A(s)), synonymous transversions (B(s)), and nonsynonymous substitutions (K(a)) into the P1/capsid region (2,643 nucleotides). Rates were determined over a 10-year period by analysis of sequences of 31 wild poliovirus type 1 isolates representing a well-defined phylogeny derived from a common imported ancestor. Similar rates were obtained by linear regression, the maximum likelihood/single-rate dated-tip method, and Bayesian inference. The very rapid K(t) [(1.03 +/- 0.10) x 10(-2) substitutions/site/year] and K(s) [(1.00 +/- 0.08) x 10(-2)] clocks were driven primarily by the A(s) clock [(0.96 +/- 0.09) x 10(-2)], the B(s) clock was approximately 10-fold slower [(0.10 +/- 0.03) x 10(-2)], and the more stochastic K(a) clock was approximately 30-fold slower [(0.03 +/- 0.01) x 10(-2)]. Nonsynonymous substitutions at all P1/capsid sites, including the neutralizing antigenic sites, appeared to be constrained by purifying selection. Simulation of the evolution of third-codon positions suggested that saturation of synonymous transitions would be evident at 10 years and complete at approximately 65 years of independent transmission. Saturation of synonymous transversions was predicted to be minimal at 20 years and incomplete at 100 years. The rapid evolution of the K(t), K(s), and A(s) clocks can be used to estimate the dates of divergence of closely related viruses, whereas the slower B(s) and K(a) clocks may be used to explore deeper evolutionary relationships within and across poliovirus genotypes.     

Pericentromeric effects shape the patterns of divergence, retention, and expression of duplicated genes in the paleopolyploid soybean

The evolutionary forces that govern the divergence and retention of duplicated genes in polyploids are poorly understood. In this study, we first investigated the rates of nonsynonymous substitution (Ka) and the rates of synonymous substitution (Ks) for a nearly complete set of genes in the paleopolyploid soybean (Glycine max) by comparing the orthologs between soybean and its progenitor species Glycine soja and then compared the patterns of gene divergence and expression between pericentromeric regions and chromosomal arms in different gene categories. Our results reveal strong associations between duplication status and Ka and gene expression levels and overall low Ks and low levels of gene expression in pericentromeric regions. It is theorized that deleterious mutations can easily accumulate in recombination-suppressed regions, because of Hill-Robertson effects. Intriguingly, the genes in pericentromeric regions-the cold spots for meiotic recombination in soybean-showed significantly lower Ka and higher levels of expression than their homoeologs in chromosomal arms. This asymmetric evolution of two members of individual whole genome duplication (WGD)-derived gene pairs, echoing the biased accumulation of singletons in pericentromeric regions, suggests that distinct genomic features between the two distinct chromatin types are important determinants shaping the patterns of divergence and retention of WGD-derived genes.     

The structure and early evolution of recently arisen gene duplicates in the Caenorhabditis elegans genome

The significance of gene duplication in provisioning raw materials for the evolution of genomic diversity is widely recognized, but the early evolutionary dynamics of duplicate genes remain obscure. To elucidate the structural characteristics of newly arisen gene duplicates at infancy and their subsequent evolutionary properties, we analyzed gene pairs with < or =10% divergence at synonymous sites within the genome of Caenorhabditis elegans. Structural heterogeneity between duplicate copies is present very early in their evolutionary history and is maintained over longer evolutionary timescales, suggesting that duplications across gene boundaries in conjunction with shuffling events have at least as much potential to contribute to long-term evolution as do fully redundant (complete) duplicates. The median duplication span of 1.4 kb falls short of the average gene length in C. elegans (2.5 kb), suggesting that partial gene duplications are frequent. Most gene duplicates reside close to the parent copy at inception, often as tandem inverted loci, and appear to disperse in the genome as they age, as a result of reduced survivorship of duplicates located in proximity to the ancestral copy. We propose that illegitimate recombination events leading to inverted duplications play a disproportionately large role in gene duplication within this genome in comparison with other mechanisms.     

Whole genome duplication of intra- and inter-chromosomes in the tomato genome

Whole genome duplication(WGD)events have been proven to occur in the evolutionary history of most angiosperms.Tomato is considered a model species of the Solanaceae family.In this study,we describe the details of the evolutionary process of the tomato genome by detecting collinearity blocks and dating the WGD events on the tree of life by combining two different methods:synonymous substitution rates(Ks)and phylogenetic trees.In total,593 collinearity blocks were discovered out of 12 pseudo-chromosomes constructed. It was evident that chromosome 2 had experienced an intra-chromosomal duplication event.Major inter-chromosomal duplication occurred among all the pseudo-chromosome.We calculated the Ks value of these collinearity blocks.Two peaks of Ks distribution were found,corresponding to two WGD events occurring approximately 36-82 million years ago(MYA)and 148-205 MYA.Additionally, the results of phylogenetic trees suggested that the more recent WGD event may have occurred after the divergence of the rosidasterid clade,but before the major diversification in Solanaceae.The older WGD event was shown to have occurred before the divergence of the rosid-asterid clade and after the divergence of rice-Arabidopsis(monocot-dicot).     

转录组测序揭示翼盖蕨(Didymochlaena trancatula)的全基因组复制历史

全基因组复制在动植物中普遍存在, 被认为是促进物种进化的重要动力之一。作为蕨类植物的单种科物种, 翼盖蕨(Didymochlaena trancatula)是真水龙骨类I的基部类群, 在蕨类中具有独特的演化地位。本研究基于高通量测序, 通过同义替换率(Ks)分析、相对定年分析揭示翼盖蕨的全基因组复制发生情况。Ks分析表明, 翼盖蕨至少经历了两次全基因组复制事件, 其中一次发生于59-62 million years ago (Mya), 另一次发生于90-94 Mya, 这两次全基因组复制事件分别和白垩纪第三纪的Cretaceous-Tertiary (C-T)大灭绝事件以及翼盖蕨的物种分化时间相吻合。进一步对两次全基因组复制保留的基因进行功能注释和富集分析, 结果显示与转录及代谢调控相关的基因优势被保留。翼盖蕨的全基因组复制事件可能促进了该物种的分化及其对极端环境的适应性。