小拟南芥MKK基因家族全基因组鉴定及进化和表达分析

丝裂原活化蛋白激酶激酶(mitogen-activated protein kinase kinase,MAPKK或MKK)是丝裂原活化蛋白激酶(mitogen-activatedproteinkinase,MAPK)级联的重要组成部分,在植物的生长发育和胁迫应答过程中发挥重要作用。目前,已在多种植物中鉴定了MKK基因家族,但在十字花科植物小拟南芥(Arabidopsis pumila)中MKK基因家族的系统鉴定与分析尚未见报道。为了探索小拟南芥MKK基因家族的进化和功能,本研究通过全基因组分析鉴定了小拟南芥中16个MKK基因,散布于小拟南芥的10条染色体上。基于系统发育分析和多重序列比对,将这些基因分为5个亚族:A亚族(5个)、B亚族(2个)、C亚族(4个)、D亚族(3个)和E亚族(2个)。分子进化和共线性分析表明小拟南芥中存在7对复制基因,分别是ApMKK1-1/1-2、ApMKK2-1/2-2、ApMKK3-1/3-2、ApMKK4-1/4-2、ApMKK5-1/5-2、ApMKK9-1/9-2和ApMKK10-1/10-2,其中ApMKK1-1/1-2在复制事件之后发生了加速进化。结合ApMKKs启动子区的顺式元件分布和ApMKKs在成熟叶片、茎、花和果实以及盐胁迫下的表达模式,结果发现复制基因的表达具有组织特异性和功能多样性。部分复制基因在组织中的表达模式存在差异,但在盐胁迫下的表达模式却基本相同。本研究结果为解析MKK介导的小拟南芥发育过程和非生物胁迫信号转导通路的复杂机制奠定了基础。     

Evolution of Stress-Regulated Gene Expression in Duplicate Genes of Arabidopsis thaliana

Due to the selection pressure imposed by highly variable environmental conditions, stress sensing and regulatory response mechanisms in plants are expected to evolve rapidly. One potential source of innovation in plant stress response mechanisms is gene duplication. In this study, we examined the evolution of stress-regulated gene expression among duplicated genes in the model plant Arabidopsis thaliana. Key to this analysis was reconstructing the putative ancestral stress regulation pattern. By comparing the expression patterns of duplicated genes with the patterns of their ancestors, duplicated genes likely lost and gained stress responses at a rapid rate initially, but the rate is close to zero when the synonymous substitution rate (a proxy for time) is >~0.8. When considering duplicated gene pairs, we found that partitioning of putative ancestral stress responses occurred more frequently compared to cases of parallel retention and loss. Furthermore, the pattern of stress response partitioning was extremely asymmetric. An analysis of putative cis-acting DNA regulatory elements in the promoters of the duplicated stress-regulated genes indicated that the asymmetric partitioning of ancestral stress responses are likely due, at least in part, to differential loss of DNA regulatory elements; the duplicated genes losing most of their stress responses were those that had lost more of the putative cis-acting elements. Finally, duplicate genes that lost most or all of the ancestral responses are more likely to have gained responses to other stresses. Therefore, the retention of duplicates that inherit few or no functions seems to be coupled to neofunctionalization. Taken together, our findings provide new insight into the patterns of evolutionary changes in gene stress responses after duplication and lay the foundation for testing the adaptive significance of stress regulatory changes under highly variable biotic and abiotic environments.     

Divergence in expression between duplicated genes in Arabidopsis

New genes may arise through tandem duplication, dispersed small-scale duplication, and polyploidy, and patterns of divergence between duplicated genes may vary among these classes. We have examined patterns of gene expression and coding sequence divergence between duplicated genes in Arabidopsis thaliana. Due to the simultaneous origin of polyploidy-derived gene pairs, we can compare covariation in the rates of expression divergence and sequence divergence within this group. Among tandem and dispersed duplicates, much of the divergence in expression profile appears to occur at or shortly after duplication. Contrary to findings from other eukaryotic systems, there is little relationship between expression divergence and synonymous substitutions, whereas there is a strong positive relationship between expression divergence and nonsynonymous substitutions. Because this pattern is pronounced among the polyploidy-derived pairs, we infer that the strength of purifying selection acting on protein sequence and expression pattern is correlated. The polyploidy-derived pairs are somewhat atypical in that they have broader expression patterns and are expressed at higher levels, suggesting differences among polyploidy- and nonpolyploidy-derived duplicates in the types of genes that revert to single copy. Finally, within many of the duplicated pairs, 1 gene is expressed at a higher level across all assayed conditions, which suggests that the subfunctionalization model for duplicate gene preservation provides, at best, only a partial explanation for the patterns of expression divergence between duplicated genes.     

Investigating ancient duplication events in the Arabidopsis genome

The complete genomic analysis of Arabidopsis thaliana has shown that a major fraction of the genome consists of paralogous genes that probably originated through one or more ancient large-scale gene or genome duplication events. However, the number and timing of these duplications still remains unclear, and several different hypotheses have been put forward recently. Here, we reanalyzed duplicated blocks found in the Arabidopsis genome described previously and determined their date of divergence based on silent substitution estimations between the paralogous genes and, where possible, by phylogenetic reconstruction. We show that methods based on averaging protein distances of heterogeneous classes of duplicated genes lead to unreliable conclusions and that a large fraction of blocks duplicated much more recently than assumed previously. We found clear evidence for one large-scale gene or even complete genome duplication event somewhere between 70 to 90 million years ago. Traces pointing to a much older (probably more than 200 million years) large-scale gene duplication event could be detected. However, for now it is impossible to conclude whether these old duplicates are the result of one or more large-scale gene duplication events. abbreviations dA, fraction of amino acid substitutions; Kn, number of nonsynonymous substitutions per nonsynonymous site; Ks, number of synonymous substitutions per synonymous site; MYA, million years ago     

Gene duplication within the Green Lineage: the case of TEL genes

Recent years have witnessed a breathtaking increase in the availability of genome sequence data, providing evidence of the highly duplicate nature of eukaryotic genomes. Plants are exceptional among eukaryotic organisms in that duplicate loci compose a large fraction of their genomes, partly because of the frequent occurrence of polyploidy (or whole-genome duplication) events. Tandem gene duplication and transposition have also contributed to the large number of duplicated genes in plant genomes. Evolutionary analyses allowed the dynamics of duplicate gene evolution to be studied and several models were proposed. It seems that, over time, many duplicated genes were lost and some of those that were retained gained new functions and/or expression patterns (neofunctionalization) or subdivided their functions and/or expression patterns between them (subfunctionalization). Recent studies have provided examples of genes that originated by duplication with successive diversification within plants. In this review, we focused on the TEL (TERMINAL EAR1-like) genes to illustrate such mechanisms. Emerged from the mei2 gene family, these TEL genes are likely to be land plant-specific. Phylogenetic analyses revealed one or two TEL copies per diploid genome. TEL gene degeneration and loss in several Angiosperm species such as in poplar and maize seem to have occurred. In Arabidopsis thaliana, whose genome experienced at least three polyploidy events followed by massive gene loss and genomic reorganization, two TEL genes were retained and two new shorter TEL-like (MCT) genes emerged. Molecular and expression analyses suggest for these genes sub- and neofunctionalization events, but confirmation will come from their functional characterization.     

Importance of lineage-specific expansion of plant tandem duplicates in the adaptive response to environmental stimuli

Plants have substantially higher gene duplication rates compared with most other eukaryotes. These plant gene duplicates are mostly derived from whole genome and/or tandem duplications. Earlier studies have shown that a large number of duplicate genes are retained over a long evolutionary time, and there is a clear functional bias in retention. However, the influence of duplication mechanism, particularly tandem duplication, on duplicate retention has not been thoroughly investigated. We have defined orthologous groups (OGs) between Arabidopsis (Arabidopsis thaliana) and three other land plants to examine the functional bias of retained duplicate genes during vascular plant evolution. Based on analysis of Gene Ontology categories, it is clear that genes in OGs that expanded via tandem duplication tend to be involved in responses to environmental stimuli, while those that expanded via nontandem mechanisms tend to have intracellular regulatory roles. Using Arabidopsis stress expression data, we further demonstrated that tandem duplicates in expanded OGs are significantly enriched in genes that are up-regulated by biotic stress conditions. In addition, tandem duplication of genes in an OG tends to be highly asymmetric. That is, expansion of OGs with tandem genes in one organismal lineage tends to be coupled with losses in the other. This is consistent with the notion that these tandem genes have experienced lineage-specific selection. In contrast, OGs with genes duplicated via nontandem mechanisms tend to experience convergent expansion, in which similar numbers of genes are gained in parallel. Our study demonstrates that the expansion of gene families and the retention of duplicates in plants exhibit substantial functional biases that are strongly influenced by the mechanism of duplication. In particular, genes involved in stress responses have an elevated probability of retention in a single-lineage fashion following tandem duplication, suggesting that these tandem duplicates are likely important for adaptive evolution to rapidly changing environments.     

Analysis of sequence, map position, and gene expression reveals conserved essential genes for iron uptake in Arabidopsis and tomato

Arabidopsis (Arabidopsis thaliana) and tomato (Lycopersicon esculentum) show similar physiological responses to iron deficiency, suggesting that homologous genes are involved. Essential gene functions are generally considered to be carried out by orthologs that have remained conserved in sequence and map position in evolutionarily related species. This assumption has not yet been proven for plant genomes that underwent large genome rearrangements. We addressed this question in an attempt to deduce functional gene pairs for iron reduction, iron transport, and iron regulation between Arabidopsis and tomato. Iron uptake processes are essential for plant growth. We investigated iron uptake gene pairs from tomato and Arabidopsis, namely sequence, conserved gene content of the regions containing iron uptake homologs based on conserved orthologous set marker analysis, gene expression patterns, and, in two cases, genetic data. Compared to tomato, the Arabidopsis genome revealed more and larger gene families coding for the iron uptake functions. The number of possible homologous pairs was reduced if functional expression data were taken into account in addition to sequence and map position. We predict novel homologous as well as partially redundant functions of ferric reductase-like and iron-regulated transporter-like genes in Arabidopsis and tomato. Arabidopsis nicotianamine synthase genes encode a partially redundant family. In this study, Arabidopsis gene redundancy generally reflected the presumed genome duplication structure. In some cases, statistical analysis of conserved gene regions between tomato and Arabidopsis suggested a common evolutionary origin. Although involvement of conserved genes in iron uptake was found, these essential genes seem to be of paralogous rather than orthologous origin in tomato and Arabidopsis.     

Expression pattern shifts following duplication indicative of subfunctionalization and neofunctionalization in regulatory genes of Arabidopsis

Gene duplication plays an important role in the evolution of diversity and novel function and is especially prevalent in the nuclear genomes of flowering plants. Duplicate genes may be maintained through subfunctionalization and neofunctionalization at the level of expression or coding sequence. In order to test the hypothesis that duplicated regulatory genes will be differentially expressed in a specific manner indicative of regulatory subfunctionalization and/or neofunctionalization, we examined expression pattern shifts in duplicated regulatory genes in Arabidopsis. A two-way analysis of variance was performed on expression data for 280 phylogenetically identified paralogous pairs. Expression data were extracted from global expression profiles for wild-type root, stem, leaf, developing inflorescence, nearly mature flower buds, and seedpod. Gene, organ, and gene by organ interaction (G x O) effects were examined. Results indicate that 85% of the paralogous pairs exhibited a significant G x O effect indicative of regulatory subfunctionalization and/or neofunctionalization. A significant G x O effect was associated with complementary expression patterns in 45% of pairwise comparisons. No association was detected between a G x O effect and a relaxed evolutionary constraint as detected by the ratio of nonsynonymous to synonymous substitutions. Ancestral gene expression patterns inferred across a Type II MADS-box gene phylogeny suggest several cases of regulatory neofunctionalization and organ-specific nonfunctionalization. Complete linkage clustering of gene expression levels across organs suggests that regulatory modules for each organ are independent or ancestral genes had limited expression. We propose a new classification, regulatory hypofunctionalization, for an overall decrease in expression level in one member of a paralogous pair while still having a significant G x O effect. We conclude that expression divergence specifically indicative of subfunctionalization and/or neofunctionalization contributes to the maintenance of most if not all duplicated regulatory genes in Arabidopsis and hypothesize that this results in increasing expression diversity or specificity of regulatory genes after each round of duplication.     

Genome-wide and molecular evolution analyses of the KT/HAK/KUP family in tomato (Solanum lycopersicum L.)

Potassium transporters belonging to the KT/HAK/KUP family play an important role in plant growth, development, mineral nutrition, and stress adaptation. In this study, we identified 19 KT/HAK/KUP family genes in tomato, distributed on 10 chromosomes, by using bioinformatics methods. A complete overview of the KT/HAK/KUP (SlHAK) genes in tomato is presented, including chromosome location, phylogeny, gene structure, and evolution pattern. Phylogenetic analysis of 19 SlHAK proteins suggested that group IV of the KT/HAK/KUP family is absent in the tomato genome. In addition, five pairs of segmental duplicated paralogs and two pairs of tandem duplicated paralogs were identified in the tomato KT/HAK/KUP family. This suggests that segmental duplication is predominant for the expansion of the SlHAK genes. Calculation of the approximate dates of duplication events using the synonymous substitution rate indicated that the segmental duplication of the KT/HAK/KUP genes in tomato originated 35.89–62.77 million years ago. Adaptive evolution analysis showed that purifying selection contributed to the evolution of segmental duplicated pairs. Furthermore, Tajima’s relative rate test indicated that all segmental duplicated pairs evolved at similar rates. As a first step toward a genome-wide analysis of the KT/HAK/KUP gene family in tomato, our results provide valuable information for understanding the function and evolution of the KT/HAK/KUP gene family in tomato and other species.     

Selection-Driven Divergence After Gene Duplication in Arabidopsis thaliana

Gene duplications are one of the most important mechanisms for the origin of evolutionary novelties. Even though various models of the fate of duplicated genes have been established, current knowledge about the role of divergent selection after gene duplication is rather limited. In this study, we analyzed sequence divergence in response to neo- and subfunctionalization of segmentally duplicated genes in the genome of Arabidopsis thaliana. We compared the genomes of A. thaliana and the poplar Populus trichocarpa to identify orthologous pairs of genes and their corresponding inparalogs. Maximum-likelihood analyses of the nonsynonymous and synonymous substitution rate ratio [Formula: see text] of pairs of A. thaliana inparalogs were used to detect differences in the evolutionary rates of protein coding sequences. We analyzed 1,924 A. thaliana paralogous pairs and our results indicate that around 6.9% show divergent ω values between the lineages for a fraction of sites. We observe an enrichment of regulatory sequences, a reduced level of co-expression and an increased number of substitutions that can be attributed to positive selection based on an McDonald-Kreitman type of analysis. Taken together, these results show that selection after duplication contributes substantially to gene novelties and hence functional divergence in plants.     

Updated map of duplicated regions in the yeast genome

We have updated the map of duplicated chromosomal segments in the Saccharomyces cerevisiae genome originally published by Wolfe and Shields in 1997 (Nature 387, 708-713). The new analysis is based on the more sensitive Smith Waterman search method instead of BLAST. The parameters used to identify duplicated chromosomal regions were optimized such as to maximize the amount of the genome placed into paired regions, under the assumption that the hypothesis that the entire genome was duplicated in a single event is correct. The core of the new map, with 52 pairs of regions containing three or more duplicated genes, is largely unchanged from our original map. 39 tRNA gene pairs and one snRNA pair have been added. To find additional pairs of genes that may have been formed by whole genome duplication, we searched through the parts of the genome that are not covered by this core map, looking for putative duplicated chromosomal regions containing only two duplicate genes instead of three, or having lower-scoring gene pairs. This approach identified a further 32 candidate paired regions, bringing the total number of protein-coding genes on the duplication map to 905 (16% of the proteome). The updated map suggests that a second copy of the ribosomal DNA array has been deleted from chromosome IV.     

Widespread paleopolyploidy in model plant species inferred from age distributions of duplicate genes

It is often anticipated that many of today's diploid plant species are in fact paleopolyploids. Given that an ancient large-scale duplication will result in an excess of relatively old duplicated genes with similar ages, we analyzed the timing of duplication of pairs of paralogous genes in 14 model plant species. Using EST contigs (unigenes), we identified pairs of paralogous genes in each species and used the level of synonymous nucleotide substitution to estimate the relative ages of gene duplication. For nine of the investigated species (wheat [Triticum aestivum], maize [Zea mays], tetraploid cotton [Gossypium hirsutum], diploid cotton [G. arboretum], tomato [Lycopersicon esculentum], potato [Solanum tuberosum], soybean [Glycine max], barrel medic [Medicago truncatula], and Arabidopsis thaliana), the age distributions of duplicated genes contain peaks corresponding to short evolutionary periods during which large numbers of duplicated genes were accumulated. Large-scale duplications (polyploidy or aneuploidy) are strongly suspected to be the cause of these temporal peaks of gene duplication. However, the unusual age profile of tandem gene duplications in Arabidopsis indicates that other scenarios, such as variation in the rate at which duplicated genes are deleted, must also be considered.     

Factors that contribute to variation in evolutionary rate among Arabidopsis genes

Surprisingly, few studies have described evolutionary rate variation among plant nuclear genes, with little investigation of the causes of rate variation. Here, we describe evolutionary rates for 11,492 ortholog pairs between Arabidopsis thaliana and A. lyrata and investigate possible contributors to rate variation among these genes. Rates of evolution at synonymous sites vary along chromosomes, suggesting that mutation rates vary on genomic scales, perhaps as a function of recombination rate. Rates of evolution at nonsynonymous sites correlate most strongly with expression patterns, but they also vary as to whether a gene is duplicated and retained after a whole-genome duplication (WGD) event. WGD genes evolve more slowly, on average, than nonduplicated genes and non-WGD duplicates. We hypothesize that levels and patterns of expression are not only the major determinants that explain nonsynonymous rate variation among genes but also a critical determinant of gene retention after duplication.     

The temporal distribution of gene duplication events in a set of highly conserved human gene families

Using a data set of protein translations associated with map positions in the human genome, we identified 1520 mapped highly conserved gene families. By comparing sharing of families between genomic windows, we identified 92 potentially duplicated blocks in the human genome containing 422 duplicated members of these families. Using branching order in the phylogenetic trees, we timed gene duplication events in these families relative to the primate-rodent divergence, the amniote-amphibian divergence, and the deuterostome-protostome divergence. The results showed similar patterns of gene duplication times within duplicated blocks and outside duplicated blocks. Both within and outside duplicated blocks, numerous duplications were timed prior to the deuterostome-protostome divergence, whereas others occurred after the amniote-amphibian divergence. Thus, neither gene duplication in general nor duplication of genomic blocks could be attributed entirely to polyploidization early in vertebrate history. The strongest signal in the data was a tendency for intrachromosomal duplications to be more recent than interchromosomal duplications, consistent with a model whereby tandem duplication-whether of single genes or of genomic blocks-may be followed by eventual separation of duplicates due to chromosomal rearrangements. The rate of separation of tandemly duplicated gene pairs onto separated chromosomes in the human lineage was estimated at 1.7 x 10(-9) per gene-pair per year.