Serial number tagging reveals a prominent sequence preference of retrotransposon integration.
Bottom Line: To address this problem we developed the serial number system, a TE tagging method that measures the frequency of integration at single nucleotide positions.We sequenced 1 million insertions of retrotransposon Tf1 in the genome of Schizosaccharomyces pombe and obtained the first profile of integration with frequencies for each individual position.Integration levels at individual nucleotides varied over two orders of magnitude and revealed that sequence recognition plays a key role in positioning integration.
Affiliation: Section on Eukaryotic Transposable Elements, Program in Cellular Regulation and Metabolism, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA.Show MeSH
Related in: MedlinePlus
Mentions: The bulk of integration sites had modest to low levels of sequence specificity (Figure 7, bit scores <0.1) suggesting that the overall pattern of integration positions was not the result of nucleotide preferences. However, we wondered whether the high numbers of independent insertions found at the ‘hottest’ positions might result from the recognition of specific nucleotides. To test this possibility we aligned the 50 insertion sites from each collection of Tf1s-neo that had the highest number of independent insertions. These 150 positions had numbers of independent insertions ranging between 71 and 622. The logo pattern from these top positions possessed a marked increase in nucleotide specificity with bit scores that in some positions were five times higher than the scores of the complete set of insertions (Figure 8A versus Figure 7A). The nucleotide preferences of Tf1 lacking the chromodomain (Tf1s-CHD-neo), at the 150 positions with the highest number of insertions also had greatly increased nucleotide specificity compared to all Tf1s-CHD-neo insertions (Figure 8B versus Figure 7B). However, the logo pattern of the top Tf1s-CHD-neo sites had nucleotide specificities higher even than the top sites of wild-type Tf1 (Figure 8B versus Figure 8A). For example, at position 18 of the top Tf1s-CHD-neo sites, the bit score was nearly 1 because 62% of the sites had a C at this location (Figure 8B and Table 3). The preference for C at position 28 was also higher in the top Tf1s-CHD-neo sites than in the top Tf1s-neo sites (Table 3, 58% versus 43%). In addition to its heightened level of specificity, Tf1 lacking the chromodomain integrated at its top 150 sites with a unique asymmetry (Figure 8B). The strongest positions of nucleotide preference only occurred downstream of the insertion sites. This surprising absence of palindromic symmetry indicates that the chromodomain influences the orientation of integration events and the recognition of nucleotides at the insertion sites with the highest number or repeated events.
Affiliation: Section on Eukaryotic Transposable Elements, Program in Cellular Regulation and Metabolism, Eunice Kennedy Shriver National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892, USA.