Diversity of LTR-retrotransposons and <Emphasis Type="Italic">Enhancer/Suppressor</Emphasis><Emphasis Type="Italic">Mutator</Emphasis>-like transposons in cassava (<Emphasis Type="Italic">Manihot esculenta</Emphasis> Crantz)

Cassava (Manihot esculenta Crantz), though a major world crop with enormous potential, is very under studied. Little is known about its genome structure and organisation. Transposable elements have a key role in the evolution of genome structure, and can be used as important tools in applied genetics. This paper sets out to survey the diversity of members of three major classes of transposable element within the cassava genome and in relation to similar elements in other plants. Members of two classes of LTR-retrotransposons, Ty1/copia-like and Ty3/gypsy-like, and of Enhancer/Suppressor Mutator (En/Spm)-like transposons were isolated and characterised. Analyses revealed 59 families of Ty1/copia, 26 families of Ty3/gypsy retrotransposons, and 40 families of En/Spm in the cassava genome. In the comparative analyses, the predicted amino acid sequences for these transposon classes were compared with those of related elements from other plant species. These revealed that there were multiple lineages of Ty1/copia-like retrotransposons in the genome of cassava and suggested that vertical and horizontal transmission as the source of cassava Mecops may not be mutually exclusive. For the Ty3/gypsy elements network, two groups of cassava Megyps were evident including the Arabidopsis Athila lineage. However, cassava En/Spm-like elements (Meens) constituted a single group within a network of plant En/Spm-like elements. Hybridisation analysis supported the presence of transposons in the genome of cassava in medium (Ty3/gypsy and En/Spm) to high (Ty1/copia) copy numbers. Thus the cassava genome was shown to contain diverse members of three major classes of transposable element; however, the different classes exhibited contrasting evolutionary histories.     

Isolation and Characterization of Ty1/copia-like Reverse Transcriptase Sequences from Mung Bean (Vigna radiata)

Ty1/copia-like sequences were amplified from mung bean (Vigna radiata (L.) Wilczek) genomic DNA, by PCR with degenerate oligonucleotide primers corresponding to highly conserved domains in the Ty1/copia-like retrotransposons. PCR fragments of roughly 270 bp were isolated and cloned, and forty clones were sequenced. Thirty-six of the forty clones had unique nucleotide sequences, and eighteen clones had a frameshift, a stop codon, or both. Alignment of the nucleotide sequences indicated that these clones, denoted Tvr, fell into nine subgroups and nine ungrouped sequences. The nucleotide sequence similarity between these elements ranged from 8% to 100%, which indicates high level of sequence heterogeneity among these clones. A phylogenetic analysis comparing these clones with corresponding sequences from other plant species showed that some of the Tvr clones are more closely related to Ty1/copia-like retrotransposons from other species than to other Tvr clones. Dot blot analysis revealed that Ty1/copia-like retrotransposons comprise about 9.3% of the mung bean genome.     

绿豆Ty1／Copia类反转录转座子逆转录酶序列的克隆和分析

利用Ty1/copia类反转录转座子的保守位点设计简并引物,从绿豆(Vignaradiata(L.)Wilczek)基因组中扩增得到了反转录转座子的逆转录酶序列。对扩增得到的约270bp的片段进行分离和克隆,并随机挑选了40个克隆进行测序,结果得到了36个单独的核酸序列,其中18个含有移码突变或终止子。根据序列比对,这些克隆可分为9组以及单个的9种。这40个克隆中,核酸序列相似性从8%到100%,显示出其核酸序列的高度异质性。将这些克隆的核酸序列与来自其他种植物的相应序列进行谱系分析,发现有些克隆与来自其他种植物的相应序列的亲缘关系比这些克隆之间更为接近。斑点杂交显示Ty1/copia反转录转座子约占绿豆基因组的9.3%。     

Translational recoding signals between gag and pol in diverse LTR retrotransposons

Because of their compact genomes, retroelements (including retrotransposons and retroviruses) employ a variety of translational recoding mechanisms to express Gag and Pol. To assess the diversity of recoding strategies, we surveyed gag/pol gene organization among retroelements from diverse host species, including elements exhaustively recovered from the genome sequences of Caenorhabditis elegans, Drosophila melanogaster, Schizosaccharomyces pombe, Candida albicans, and Arabidopsis thaliana. In contrast to the retroviruses, which typically encode pol in the -1 frame relative to gag, nearly half of the retroelements surveyed encode a single gag-pol open reading frame. This was particularly true for the Ty1/copia group retroelements. Most animal Ty3/gypsy retroelements, on the other hand, encode gag and pol in separate reading frames, and likely express Pol through +1 or -1 frameshifting. Conserved sequences conforming to slippery sites that specify viral ribosomal frameshifting were identified among retroelements with pol in the -1 frame. None of the plant retroelements encoded pol in the -1 frame relative to gag; however, two closely related plant Ty3/gypsy elements encode pol in the +1 frame. Interestingly, a group of plant Ty1/copia retroelements encode pol either in a +1 frame relative to gag or in two nonoverlapping reading frames. These retroelements have a conserved stem-loop at the end of gag, and likely express pol either by a novel means of internal ribosomal entry or by a bypass mechanism.     

Retrotransposon populations of <Emphasis Type="Italic">Vicia</Emphasis> species with varying genome size

The (non-LTR) LINE and Ty3-gypsy-type LTR retrotransposon populations of three Vicia species that differ in genome size (Vicia faba, Vicia melanops and Vicia sativa) have been characterised. In each species the LINE retrotransposons comprise a complex, very heterogeneous set of sequences, while the Ty3-gypsy elements are much more homogeneous. Copy numbers of all three retrotransposon groups (Ty1-copia, Ty3-gypsy and LINE) in these species have been estimated by random genomic sequencing and Southern hybridisation analysis. The Ty3-gypsy elements are extremely numerous in all species, accounting for 18–35% of their genomes. The Ty1-copia group elements are somewhat less abundant and LINE elements are present in still lower amounts. Collectively, 20–45% of the genomes of these three Vicia species are comprised of retrotransposons. These data show that the three retrotransposon groups have proliferated to different extents in members of the Vicia genus and high proliferation has been associated with homogenisation of the retrotransposon population.Electronic Supplementary Material Supplementary material is available for this article at .