The core domain of retrotransposon integrase in Hordeum: predicted structure and evolution

Propagation of long terminal repeat (LTR)-bearing retrotransposons and retroviruses requires integrase (IN, EC 2.7.7.-), encoded by the retroelements themselves, which mediates the insertion of cDNA copies back into the genome. An active retrotransposon family, BARE-1, comprises approximately 7% of the barley (Hordeum vulgare subsp. vulgare) genome. We have generated models for the secondary and tertiary structure of BARE-1 IN and demonstrate their similarity to structures for human immunodeficiency virus 1 and avian sarcoma virus INs. The IN core domains were compared for 80 clones from 28 Hordeum accessions representative of the diversity of the genus. Based on the structural model, variations in the predicted, aligned translations from these clones would have minimal structural and functional effects on the encoded enzymes. This indicates that Hordeum retrotransposon IN has been under purifying selection to maintain a structure typical of retroviral INs. These represent the first such analyses for plant INs.      

A centromeric tandem repeat family originating from a part of Ty3/gypsy-retroelement in wheat and its relatives

From a wild diploid species that is a relative of wheat, Aegilops speltoides, a 301-bp repeat containing 16 copies of a CAA microsatellite was isolated. Southern blot and fluorescence in situ hybridization revealed that approximately 250 bp of the sequence is tandemly arrayed at the centromere regions of A- and B-genome chromosomes of common wheat and rye chromosomes. Although the DNA sequence of this 250-bp repeat showed no notable homology in the databases, the flanking or intervening sequences between the repeats showed high homologies (>82%) to two separate sequences of the gag gene and its upstream region in cereba, a Ty3/gypsy-like retroelement of Hordeum vulgare. Since the amino acid sequence deduced from the 250 bp with seven CAAs showed some similarity ( approximately 53%) to that of the gag gene, we concluded that the 250-bp repeats had also originated from the cereba-like retroelements in diploid wheat such as Ae. speltoides and had formed tandem arrays, whereas the 300-bp repeats were dispersed as a part of cereba-like retroelements. This suggests that some tandem repeats localized at the centromeric regions of cereals and other plant species originated from parts of retrotransposons.     

Genomic organization, evolution, and structural peculiarities of highly repetitive DNA of Hordeum vulgare

A fraction of highly repeated DNA sequences of Hordeum vulgare has been investigated by cloning 19 separate highly repetitive sequences in the plasmid pBR327. Characteristics studied included genus specificity of isolated sequences, their prevalence, and genome organization. Sequences (pHv7161, pHv7191, pHv7179) have been identified that are the most widespread in the H. vulgare genome and have a complicated arrangement. A tandemly arranged sequence, pHv7141, was also identified. The primary structure of a 999 bp long, BamHI fragment of one of the most widespread sequences, pHv7161, as well as the adjacent pHv7302 and pHv7245 sequences was determined. The fragment abounds in inverted repeats, of which two are flanked by direct repeats, and contains short subrepeats, A, B, and C, and a great variety of potential protein-binding sites. A comparison is drawn between the content and genome organization of highly repeated DNA sequences of H. vulgare and those of the wild barley species Hordeum bulbosum, Hordeum jubatum, Hordeum geniculatum, Hordeum brevisubulatum, Hordeum turkestanicum, and Hordeum murinum. According to the above characters (close copy number and genome organization similarity of highly repetitive sequences) the species under discussion have been classified into four groups. This division is in good agreement with other data on interspecific crossing in Hordeum and on chromosome pairing in hybrid meiosis.     

Interspecies variability in the organization of repeated sequences of the genus Hordeum

Seven barley species have been compared for organization of repeated sequences. Quantitative variation of repeated DNA fractions is demonstrated, though the total amount of sequences (reassociation up to Cot=10) in most cases does not vary. The repeats are divided into four groups by the mode of interspecific variability, with the help of dot and blot hybridization of the genomes under study with cloned highly repeated sequences of Hordeum vulgare. The first group contains the pHv7161 family of the most conservative sequences. The second group comprises moderately changing repeats. The third group includes highly variable Hind III repeats of Hordeum genomes, and the fourth group is represented by pHv7191 family of repeats that are highly amplified in H. vulgare genome. Comparative analysis of content and organization of highly repeated sequences in genome helps to clarify phylogenetic relationships in the genus and can be used for prediction of successfullness of interspecific hybridization.     

Phylogeny in the genus Hordeum based on nucleotide sequences closely linked to the vrs1 locus (row number of spikelets).

The phylogenetic relationship between four basic genomes designated H, I, Xa, and Xu in the genus Hordeum was studied using a nuclear DNA sequence. The sequence, cMWG699, is single copy in the H. vulgare genome, and tightly linked to the vrs1 locus which controls two- and six-rowed spikes. DNA fragments homologous to cMWG699 were amplified from diploid Hordeum species and the nucleotide sequences were determined. A phylogeny based on both base substitutions and an insertion-deletion event showed that the H- and Xa-genome groups are positioned in one monophyletic group indicating that the Xa-genome taxa should be included in the H-genome group. The large H-genome group is highly homogeneous. The I and Xu genomes are distinctly separated from H and Xa, and form sister groups. Another phylogeny pattern based on data excluding the insertion-deletion gave a result that the Xa genome forms a sister group to the H-genome group. The difference between the H and Xa genomes was affected only by a single base insertion-deletion event, thus the H and Xa genomes are likely to be closely related. The I and Xu genomes were again distinctly separated from the H and Xa genomes.     

Survey of long terminal repeat retrotransposons of domesticated silkworm (Bombyx mori)

Long terminal retrotransposons are major components of eukaryotic transposable elements. We have surveyed the long terminal repeats (LTR) retrotransposons of domesticated silkworm (Bombyx mori) by mining the data produced by Bombyx mori Genome Sequencing Project. At least 29 separate families of LTR retrotransposons are identified in this survey, comprising of 11.8% of the complete sequence. Families of domesticated silkworm LTR retrotransposons can be mainly classified into three groups: gypsy-like, copia-like, Pao-Bel. Fourteen families identified consist of gypsy-like elements, four families consist of copia-like elements and seven families consist of Pao-Bel elements. In addition to the three groups of LTR retrotransposons, two families of unusual non-coding elements are identified in the genome of this species. Further phylogenetic analysis of RT domain indicates that the elements of B.mori show high diversity and can form different clades in each group. An analysis of sequence variation from different families reveals distinct patterns of variation for the elements belonging to three groups. The analysis of the domesticated silkworm LTR retrotransposons should assist in our understanding of the roles of retroelement in lepidopteron insect genome evolution.     

Non-LTR retrotransposons in fungi

Non-long terminal repeat (non-LTR) retrotransposons have contributed to shaping the structure and function of genomes. Fungi have small genomes, usually with limited amounts of repetitive DNA. In silico approach has been used to survey the non-LTR elements in 57 fungal genomes. More than 100 novel non-LTR retrotransposons were found, which belonged to five diverse clades. The present survey identified two novel clades of fungal non-LTR retrotransposons. The copy number of non-LTR retroelements varied widely. Some of the studied species contained a single copy of non-LTR retrotransposon, whereas others possessed a great number of non-LTR retrotransposon copies per genome. Although evolutionary relationships of most elements are congruent with phylogeny of host species, a new case of possible horizontal transfer was found between Eurotiomycetes and Sordariomycetes. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Comparative sequence analysis of colinear barley and rice bacterial artificial chromosomes

Colinearity of a large region from barley (Hordeum vulgare) chromosome 5H and rice (Oryza sativa) chromosome 3 has been demonstrated by mapping of several common restriction fragment-length polymorphism clones on both regions. One of these clones, WG644, was hybridized to rice and barley bacterial artificial chromosome (BAC) libraries to select homologous clones. One BAC from each species with the largest overlapping segment was selected by fingerprinting and blot hybridization with three additional restriction fragment-length polymorphism clones. The complete barley BAC 635P2 and a 50-kb segment of the rice BAC 36I5 were completely sequenced. A comparison of the rice and barley DNA sequences revealed the presence of four conserved regions, containing four predicted genes. The four genes are in the same orientation in rice, but the second gene is in inverted orientation in barley. The fourth gene is duplicated in tandem in barley but not in rice. Comparison of the homeologous barley and rice sequences assisted the gene identification process and helped determine individual gene structures. General gene structure (exon number, size, and location) was largely conserved between rice and barley and to a lesser extent with homologous genes in Arabidopsis. Colinearity of these four genes is not conserved in Arabidopsis compared with the two grass species. Extensive similarity was not found between the rice and barley sequences other than within the exons of the structural genes, and short stretches of homology in the promoters and 3' untranslated regions. The larger distances between the first three genes in barley compared with rice are explained by the insertion of different transposable retroelements.     

Phylogenetic determination of the pace of transposable element proliferation in plants: copia and LINE-like elements in Gossypium.

Transposable elements contribute significantly to plant genome evolution in myriad ways, ranging from local insertional mutations to global effects exerted on genome size through accumulation. Differential accumulation and deletion of transposable elements may profoundly affect genome size, even among members of the same genus. One example is that of Gossypium (cotton), where much of the 3-fold genome size variation is due to differential accumulation of one gypsy-like LTR retrotransposon, Gorge3. Copia and non-LTR LINE retrotransposons are also major components of the Gossypium genome, but unlike Gorge3, their extant copy numbers do not correlate with genome size. In the present study, we describe the nature and timing of transposition for copia and LINE retrotransposons in Gossypium. Our findings indicate that copia retrotransposons have been active in each lineage since divergence from a common ancestor, and that they have proliferated in a punctuated manner. However, the evolutionary history of LINEs contrasts markedly with that of the copia retrotransposons. Although LINEs have also been active in each lineage, they have accumulated in a stochastically regular manner, and phylogenetic analysis suggests that extant LINE populations in Gossypium are dominated by ancient insertions. Interestingly, the magnitude of transpositional bursts in each lineage corresponds directly with extant estimated copy number.