Comparative DNA sequence analysis of mapped wheat ESTs reveals the complexity of genome relationships between rice and wheat

The use of DNA sequence-based comparative genomics for evolutionary studies and for transferring information from model species to related large-genome species has revolutionized molecular genetics and breeding strategies for improving those crops. Comparative sequence analysis methods can be used to cross-reference genes between species maps, enhance the resolution of comparative maps, study patterns of gene evolution, identify conserved regions of the genomes, and facilitate interspecies gene cloning. In this study, 5,780 Triticeae ESTs that have been physically mapped using wheat (Triticum aestivum L.) deletion lines and segregating populations were compared using NCBI BLASTN to the first draft of the public rice (Oryza sativa L.) genome sequence data from 3,280 ordered BAC/PAC clones. A rice genome view of the homoeologous wheat genome locations based on sequence analysis shows general similarity to the previously published comparative maps based on Southern analysis of RFLP. For most rice chromosomes there is a preponderance of wheat genes from one or two wheat chromosomes. The physical locations of non-conserved regions were not consistent across rice chromosomes. Some wheat ESTs with multiple wheat genome locations are associated with the non-conserved regions of similarity between rice and wheat. The inverse view, showing the relationship between the wheat deletion map and rice genomic sequence, revealed the breakdown of gene content and order at the resolution conferred by the physical chromosome deletions in the wheat genome. An average of 35% of the putative single copy genes that were mapped to the most conserved bins matched rice chromosomes other than the one that was most similar. This suggests that there has been an abundance of rearrangements, insertions, deletions, and duplications eroding the wheat-rice genome relationship that may complicate the use of rice as a model for cross-species transfer of information in non-conserved regions.     

Signals of Historical Interlocus Gene Conversion in Human Segmental Duplications

Standard methods of DNA sequence analysis assume that sequences evolve independently, yet this assumption may not be appropriate for segmental duplications that exchange variants via interlocus gene conversion (IGC). Here, we use high quality multiple sequence alignments from well-annotated segmental duplications to systematically identify IGC signals in the human reference genome. Our analysis combines two complementary methods: (i) a paralog quartet method that uses DNA sequence simulations to identify a statistical excess of sites consistent with inter-paralog exchange, and (ii) the alignment-based method implemented in the GENECONV program. One-quarter (25.4%) of the paralog families in our analysis harbor clear IGC signals by the quartet approach. Using GENECONV, we identify 1477 gene conversion tracks that cumulatively span 1.54 Mb of the genome. Our analyses confirm the previously reported high rates of IGC in subtelomeric regions and Y-chromosome palindromes, and identify multiple novel IGC hotspots, including the pregnancy specific glycoproteins and the neuroblastoma breakpoint gene families. Although the duplication history of a paralog family is described by a single tree, we show that IGC has introduced incredible site-to-site variation in the evolutionary relationships among paralogs in the human genome. Our findings indicate that IGC has left significant footprints in patterns of sequence diversity across segmental duplications in the human genome, out-pacing the contributions of single base mutation by orders of magnitude. Collectively, the IGC signals we report comprise a catalog that will provide a critical reference for interpreting observed patterns of DNA sequence variation across duplicated genomic regions, including targets of recent adaptive evolution in humans.     

Patterns of Gene Duplication and Their Contribution to Expansion of Gene Families in Grapevine

Grapevine is an important fruit crop that has undergone a long history of evolution. Analysis of the whole genome sequence of grapevine has revealed presence of an early palaeo-hexaploid along with three complements. Thus, gene duplication and genome expansion are common in this genome. In this study, we identified 17,922 duplicated genes in the whole grapevine genome. Among these, 2,039; 628; 1,428; 722; and 2,942 were identified respectively as produced by genome-wide, tandem, proximal, retrotransposed, and DNA-based transposed duplications. Analyses of the evolutionary patterns for different types of duplication using non-synonymous and synonymous substitution rates uncovered a series of underlying rules. Thereafter, all the grapevine genes were classified into families, and the contributions of different types of duplication to the expansion of large families were revealed. No duplication type was solely responsible for the formation of any large gene family, but some families showed enrichment of a special type of duplication. On the basis of this study, we believe that uncovering the underlying rules for gene duplications, expansions of gene families, and their evolutionary styles will contribute significantly to a comprehensive understanding of the features of the grapevine genome.     

Characterization of paralogous protein families in rice

Background  

High gene numbers in plant genomes reflect polyploidy and major gene duplication events. Oryza sativa, cultivated rice, is a diploid monocotyledonous species with a ~390 Mb genome that has undergone segmental duplication of a substantial portion of its genome. This, coupled with other genetic events such as tandem duplications, has resulted in a substantial number of its genes, and resulting proteins, occurring in paralogous families.     

A comprehensive expression analysis of all members of a gene family encoding cell-wall enzymes allowed us to predict cis-regulatory regions involved in cell-wall construction in specific organs of Arabidopsis.

The Arabidopsis thaliana genome sequencing project has revealed that multigene families, such as those generated by genome duplications, are more abundant among plant genomes than among animal genomes. To gain insight into the evolutionary implications of the multigene families in higher plants, we examined the XTH gene family, a group of genes encoding xyloglucan endotransglucosylase/hydrolase, which are responsible for cell-wall construction in plants. Expression analysis of all members (33 genes) of this family, using quantitative real-time RT-PCR, revealed that most members exhibit distinct expression profiles in terms of tissue specificity and responses to hormonal signals, with some members exhibiting similar expression patterns. By comparing the flanking sequences of individual genes, we identified four sets of large-segment duplications and two sets of solitary gene duplications. In each set of gene duplicates, long nucleotide sequences, ranging from one to two hundred base pairs, are conserved. Furthermore, gene duplicates exhibit similar organ-specific expression profiles. These facts allowed us to predict putative cis-regulatory regions, particularly those responsible for cell-wall construction, and hence for morphogenesis, that are specific for certain organs or tissues in plants.     

Extensive duplications of phototransduction genes in early vertebrate evolution correlate with block (chromosome) duplications

Many gene families in mammals have members that are expressed more or less uniquely in the retina or differentially in specific retinal cell types. We describe here analyses of nine such gene families with regard to phylogenetic relationships and chromosomal location. The families are opsins, G proteins (alpha, beta, and gamma subunits), phosphodiesterases type 6, cyclic nucleotide-gated channels, G-protein-coupled receptor kinases, arrestins, and recoverins. The results suggest that multiple new gene copies arose in all of these families very early in vertebrate evolution during a period with extensive gene duplications. Many of the new genes arose through duplications of large chromosome regions (blocks of genes) or even entire chromosomes, as shown by linkage with other gene families. Some of the phototransduction families belong to the same duplicated regions and were thus duplicated simultaneously. We conclude that gene duplications in early vertebrate evolution probably helped facilitate the specialization of the retina and the subspecialization of different retinal cell types.     

A novel gene family NBPF: intricate structure generated by gene duplications during primate evolution

Partial and complete genome duplications occurred during evolution and resulted in the creation of new genes and gene families. We identified a novel and intricate human gene family located primarily in regions of segmental duplications on human chromosome 1. We named it NBPF, for neuroblastoma breakpoint family, because one of its members is disrupted by a chromosomal translocation in a neuroblastoma patient. The NBPF genes have a repetitive structure with high intragenic and intergenic sequence similarity in both coding and noncoding regions. These similarities might expose these genomic regions to illegitimate recombination, resulting in structural variation in the NBPF genes. The encoded proteins contain a highly conserved domain of unknown function, which we have named the NBPF repeat. In silico analysis combined with the isolation of multiple full-length cDNA clones showed that several members of this gene family are abundantly expressed in a large variety of tissues and cell lines. Strikingly, no discernable orthologues could be identified in the completed genomes of fruit fly, nematode, mouse, or rat, but sequences with low homology could be isolated from the draft canine and bovine genomes. Interestingly, this gene family shows primate-specific duplications that result in species-specific arrays of NBPF homologous sequences. Overall, this novel NBPF family reflects the continuous evolution of primate genomes that resulted in large physiological differences, and its potential role in this process is discussed.     

Comparative sequence analysis of the region harboring the hardness locus in barley and its colinear region in rice

The ancestral shared synteny concept has been advocated as an approach to positionally clone genes from complex genomes. However, the unified grass genome model and the study of grasses as a single syntenic genome is a topic of considerable controversy. Hence, more quantitative studies of cereal colinearity at the sequence level are required. This study compared a contiguous 300-kb sequence of the barley (Hordeum vulgare) genome with the colinear region in rice (Oryza sativa). The barley sequence harbors genes involved in endosperm texture, which may be the subject of distinctive evolutionary forces and is located at the extreme telomeric end of the short arm of chromosome 5H. Comparative sequence analysis revealed the presence of five orthologous genes and a complex, postspeciation evolutionary history involving small chromosomal rearrangements, a translocation, numerous gene duplications, and extensive transposon insertion. Discrepancies in gene content and microcolinearity indicate that caution should be exercised in the use of rice as a surrogate for map-based cloning of genes from large genome cereals such as barley.     

Rapid evolutionary dynamics in a 2.8‐Mb chromosomal region containing multiple prolamin and resistance gene families in Aegilops tauschii

Prolamin and resistance gene families are important in wheat food use and in defense against pathogen attacks, respectively. To better understand the evolution of these multi‐gene families, the DNA sequence of a 2.8‐Mb genomic region, representing an 8.8 cM genetic interval and harboring multiple prolamin and resistance‐like gene families, was analyzed in the diploid grass Aegilops tauschii, the D‐genome donor of bread wheat. Comparison with orthologous regions from rice, Brachypodium, and sorghum showed that the Ae. tauschii region has undergone dramatic changes; it has acquired more than 80 non‐syntenic genes and only 13 ancestral genes are shared among these grass species. These non‐syntenic genes, including prolamin and resistance‐like genes, originated from various genomic regions and likely moved to their present locations via sequence evolution processes involving gene duplication and translocation. Local duplication of non‐syntenic genes contributed significantly to the expansion of gene families. Our analysis indicates that the insertion of prolamin‐related genes occurred prior to the separation of the Brachypodieae and Triticeae lineages. Unlike in Brachypodium, inserted prolamin genes have rapidly evolved and expanded to encode different classes of major seed storage proteins in Triticeae species. Phylogenetic analyses also showed that the multiple insertions of resistance‐like genes and subsequent differential expansion of each R gene family. The high frequency of non‐syntenic genes and rapid local gene evolution correlate with the high recombination rate in the 2.8‐Mb region with nine‐fold higher than the genome‐wide average. Our results demonstrate complex evolutionary dynamics in this agronomically important region of Triticeae species.     

Genome Duplication in Soybean (Glycine Subgenus Soja)

Restriction fragment length polymorphism mapping data from nine populations (Glycine max X G. soja and G. max X G. max) of the Glycine subgenus soja genome led to the identification of many duplicated segments of the genome. Linkage groups contained up to 33 markers that were duplicated on other linkage groups. The size of homoeologous regions ranged from 1.5 to 106.4 cM, with an average size of 45.3 cM. We observed segments in the soybean genome that were present in as many as six copies with an average of 2.55 duplications per segment. The presence of nested duplications suggests that at least one of the original genomes may have undergone an additional round of tetraploidization. Tetraploidization, along with large internal duplications, accounts for the highly duplicated nature of the genome of the subgenus. Quantitative trait loci for seed protein and oil showed correspondence across homoeologous regions, suggesting that the genes or gene families contributing to seed composition have retained similar functions throughout the evolution of the chromosomes.     

Contrasting rates of evolution in Pm3 loci from three wheat species and rice

The Pm3 gene from wheat confers resistance against powdery mildew and recent studies have shown that it is a member of a multigene family in the wheat genome. We compared genomic sequences ranging from 178 to 332 kb containing six Pm3-like genes and five gene fragments from orthologous loci in the A genome of wheat at three different ploidy levels. We found that the wheat Pm3 loci display an extremely dynamic evolution where sequence conservation is minimal between species and basically limited to very short sequences containing the genetic markers that define the orthology. The Pm3-like genes and their up- and downstream regions were reshuffled by multiple rearrangements, resulting in a complex mosaic of conserved and unique sequences. Comparison with rice showed that the known wheat Pm3-like genes represent only one branch of a large superfamily with several clusters in rice and suggests the presence of additional similar genes in the wheat genome. Estimates of divergence times and transposable-element insertions indicate that the Pm3 locus in wheat has undergone more drastic changes in its recent evolution than its counterpart in rice. This indicates that loci containing homologous resistance gene analogs can evolve at highly variable speeds in different species.     

Evidence that rice and other cereals are ancient aneuploids

Detailed analyses of the genomes of several model organisms revealed that large-scale gene or even entire-genome duplications have played prominent roles in the evolutionary history of many eukaryotes. Recently, strong evidence has been presented that the genomic structure of the dicotyledonous model plant species Arabidopsis is the result of multiple rounds of entire-genome duplications. Here, we analyze the genome of the monocotyledonous model plant species rice, for which a draft of the genomic sequence was published recently. We show that a substantial fraction of all rice genes ( approximately 15%) are found in duplicated segments. Dating of these block duplications, their nonuniform distribution over the different rice chromosomes, and comparison with the duplication history of Arabidopsis suggest that rice is not an ancient polyploid, as suggested previously, but an ancient aneuploid that has experienced the duplication of one-or a large part of one-chromosome in its evolutionary past, approximately 70 million years ago. This date predates the divergence of most of the cereals, and relative dating by phylogenetic analysis shows that this duplication event is shared by most if not all of them.     

Conserved synteny between the Ciona genome and human paralogons identifies large duplication events in the molecular evolution of the insulin-relaxin gene family

The aims of the study were to outline the sequence of eventsthat gave rise to the vertebrate insulin-relaxin gene familyand the chromosomal regions in which they reside. We analyzedthe gene content surrounding the human insulin/relaxin geneswith respect to what family they belonged to and if the duplicationhistory of investigated families parallels the evolution ofthe insulin-relaxin family members. Markov Clustering and phylogeneticanalysis were used to determine family identity. More than 15%of the genes belonged to families that have paralogs in theregions, defining two sets of quadruplicate paralogy regions.Thereby, the localization of insulin/relaxin genes in humansis in accordance with those regions on human chromosomes 1,11, 12, 19q (insulin/insulin-like growth factors) and 1, 6p/15q,9/5, 19p (insulin-like factors/relaxins) were formed duringtwo genome duplications. We compared the human genome with thatof Ciona intestinalis, a species that split from the vertebratelineage before the two suggested genome duplications. Two insulin-likeorthologs were discovered in addition to the already describedCi-insulin gene. Conserved synteny between the Ciona regionshosting the insulin-like genes and the two sets of human paralogonsimplies their common origin. Linkage of the two human paralogons,as seen in human chromosome 1, as well as the two regions hostingthe Ciona insulin-like genes suggests that a segmental duplicationgave rise to the region prior to the genome doublings. Thus,preserved gene content provides support that genome duplication(s)in addition to segmental and single-gene duplications shapedthe genomes of extant vertebrates.     

Gene content and density in banana (<Emphasis Type="Italic">Musa acuminata</Emphasis>) as revealed by genomic sequencing of BAC clones

The complete sequence of Musa acuminata bacterial artificial chromosome (BAC) clones is presented and, consequently, the first analysis of the banana genome organization. One clone (MuH9) is 82,723 bp long with an overall G+C content of 38.2%. Twelve putative protein-coding sequences were identified, representing a gene density of one per 6.9 kb, which is slightly less than that previously reported for Arabidopsis but similar to rice. One coding sequence was identified as a partial M. acuminata malate synthase, while the remaining sequences showed a similarity to predicted or hypothetical proteins identified in genome sequence data. A second BAC clone (MuG9) is 73,268 bp long with an overall G+C content of 38.5%. Only seven putative coding regions were discovered, representing a gene density of only one gene per 10.5 kb, which is strikingly lower than that of the first BAC. One coding sequence showed significant homology to the soybean ribonucleotide reductase (large subunit). A transition point between coding regions and repeated sequences was found at approximately 45 kb, separating the coding upstream BAC end from its downstream end that mainly contained transposon-like sequences and regions similar to known repetitive sequences of M. acuminata. This gene organization resembles Gramineae genome sequences, where genes are clustered in gene-rich regions separated by gene-poor DNA containing abundant transposons.Communicated by J.S. Heslop-Harrison     

A bacterial genome in flux: the twelve linear and nine circular extrachromosomal DNAs in an infectious isolate of the Lyme disease spirochete Borrelia burgdorferi

We have determined that Borrelia burgdorferi strain B31 MI carries 21 extrachromosomal DNA elements, the largest number known for any bacterium. Among these are 12 linear and nine circular plasmids, whose sequences total 610 694 bp. We report here the nucleotide sequence of three linear and seven circular plasmids (comprising 290 546 bp) in this infectious isolate. This completes the genome sequencing project for this organism; its genome size is 1 521 419 bp (plus about 2000 bp of undetermined telomeric sequences). Analysis of the sequence implies that there has been extensive and sometimes rather recent DNA rearrangement among a number of the linear plasmids. Many of these events appear to have been mediated by recombinational processes that formed duplications. These many regions of similarity are reflected in the fact that most plasmid genes are members of one of the genome's 161 paralogous gene families; 107 of these gene families, which vary in size from two to 41 members, contain at least one plasmid gene. These rearrangements appear to have contributed to a surprisingly large number of apparently non-functional pseudogenes, a very unusual feature for a prokaryotic genome. The presence of these damaged genes suggests that some of the plasmids may be in a period of rapid evolution. The sequence predicts 535 plasmid genes >/=300 bp in length that may be intact and 167 apparently mutationally damaged and/or unexpressed genes (pseudogenes). The large majority, over 90%, of genes on these plasmids have no convincing similarity to genes outside Borrelia, suggesting that they perform specialized functions.