Characterization of ten novel Ty1/copia-like retrotransposon families of the grapevine genome

Cédric Moisy,Keith E Garrison,Frédérique Pelsy,Carole P Meredith

doi:10.1186/1471-2164-9-469

Abstract

BackgroundRetrotransposons make a significant contribution to the size, organization and genetic diversity of their host genomes. To characterize retrotransposon families in the grapevine genome (the fourth crop plant genome sequenced) we have combined two approaches: a PCR-based method for the isolation of RnaseH-LTR sequences with a computer-based sequence similarity search in the whole-genome sequence of PN40024.ResultsSupported by a phylogenic analysis, ten novel Ty1/copia families were distinguished in this study. To select a canonical reference element sequence from amongst the various insertions in the genome belonging to each retroelement family, the following screening criteria were adopted to identify the element sequence with: (1) perfect 5 bp-duplication of target sites, (2) the highest level of identity between 5' and 3'-LTR within a single insertion sequence, and (3) longest, un-interrupted coding capacity within the gag-pol ORF. One to eight copies encoding a single putatively functional gag-pol polyprotein were found for three families, indicating that these families could be still autonomous and active. For the others, no autonomous copies were identified. However, a subset of copies within the presumably non-autonomous families had perfect identity between their 5' and 3' LTRs, indicating a recent insertion event. A phylogenic study based on the sequence alignment of the region located between reverse transcriptase domains I and VII distinguished these 10 families from other plant retrotransposons. Including the previously characterized Ty1/copia-like grapevine retrotransposons Tvv1 and Vine 1 and the Ty3/gypsy-like Gret1 in this assessment, a total of 1709 copies were identified for the 13 retrotransposon families, representing 1.24% of the sequenced genome. The copy number per family ranged from 91–212 copies. We performed insertion site profiling for 8 out of the 13 retrotransposon families and confirmed multiple insertions of these elements across the Vitis genus. Insertional polymorphism analysis and dating of full-length copies based on their LTR divergence demonstrated that each family has a particular amplification history, with 71% of the identified copies being inserted within the last 2 million years.ConclusionThe strategy we used efficiently delivered new Ty1/copia-like retrotransposon sequences, increasing the total number of characterized grapevine retrotrotransposons from 3 to 13. We provide insights into the representation and dynamics of the 13 families in the genome. Our data demonstrated that each family has a particular amplification pattern, with 7 families having copies recently inserted within the last 0.2 million year. Among those 7 families with recent insertions, three retain the capacity for activity in the grape genome today.

Highlights

Retrotransposons make a significant contribution to the size, organization and genetic diversity of their host genomes
The intermediate RNA transcript is reverse-transcribed into DNA by a retrotransposon-encoded reverse transcriptase enzyme, and the DNA is inserted by additional retrotransposonencoded enzymes into the host genome at a new location, increasing the copy number of the retrotransposon family [7]
Identification of the novel retrotransposon families Combining an RnaseH-long terminal repeat (LTR) analysis of sequences isolated using the technique described by Pearce et al [25] with a computer-based sequence similarity search in the wholegenome grapevine sequence, we have identified ten novel Ty1/copia-like retrotransposon families

Summary

Introduction

Retrotransposons make a significant contribution to the size, organization and genetic diversity of their host genomes. The intermediate RNA transcript is reverse-transcribed into DNA by a retrotransposon-encoded reverse transcriptase enzyme, and the DNA is inserted by additional retrotransposonencoded enzymes into the host genome at a new location, increasing the copy number of the retrotransposon family [7]. The copy-and-paste cycle of retrotransposition in the host genome requires the synthesis of specific proteins encoded by two major retrotransposon genes, gag and pol. In Ty1/copia elements, the pol gene encodes the protease, integrase, reverse transcriptase (RT), and RnaseH domains in that order from the 5' end of the gene. The enzymatic functions encoded in each of these domains are required to synthesize an extrachromosomal DNA daughter copy by reverse-transcription from the intermediate RNA prior to its reinsertion into the genome. One hallmark of retroelements is the genetic heterogeneity of individual copies belonging to the same family [14]

Methods

Results

Discussion

Conclusion