with no matching siRNAs tend to have uniform DNA methylation density throughout their length and are typically ﬂanked by unmethylated sequences (data not shown). TE sequences and ﬂanking genes It was previously shown that TE sequences tend to be less methylated when located close to genes, presumably because of deleterious effects of TE methylation on the expression of neighboring genes (28). The proportion of unmethylated TE sequences was reported to drop from 55% within genes to below 20% for the ﬁrst 500 bp window away from genes, with little further decrease beyond this point. However, this analysis did not distin- guish euchromatic from heterochromatiic genes (28), which are characterized by dramatically distinct intergenic regions (short and TE-poor versus long and TE-rich, respectively). This prompted us to explore further the underrepresentation of methylated TE sequences near genes using our extended dataset and only considering genes within euchromatin. To this end, methylated and unmethylated TE sequences were scored in 100-bp windows for a distance of up to 1 kb upstream and Figure 8. Distance between TE sequences and genes in euchromatin. (a) Both methylated and unmethylated TE sequences tend to accumu- late close to genes. Note that because results do not substantially differ for the 50- and 30-ends of genes, they are not distinguished in the ﬁgure. (b) The proportion of unmethylated to total TE sequences drops slightly farther away from the 50-end of genes. No similar drop is observed from the 30-end of genes. Only the TE sequence closest to the start or stop codon was considered for this analysis. Nucleic Acids Research, 2011, Vol. 39, No. 16 6929 downstream of genes. Our analysis reveals that in eu- chromatin, both methylated and unmethylated TE se- quences tend in fact to over accumulate close to the 50- and 30-ends of genes (Figure 8a). Moreover, although the ratio of unmethylated versus methylated TE sequences drops with distance away from genes, as previously reported (28), this drop is rather limited (60% to a minimum of 40%), speciﬁc to the 50-end of genes and less discernible when considering only methylated TE se- quences with matching siRNAs, which are the least abundant overall (Figure 8b and Supplementary Table S2). These results suggest therefore that methylated TE sequences have more deleterious effects on transcription initiation than termination and that these effects are more severe when methylated TE sequences have matching siRNAs. We next tested if spreading of DNA methylation from siRNA-targeted TE sequences could provide a plausible explanation for the deleterious effects of TE methylation on gene expression. For this, DNA methylation densities were calculated in non-overlapping 100-bp windows for all methylated euchromatic TE sequences (n = 401) associated with 24-nt siRNAs and ﬂanked by sequences not associated with siRNAs. Although spreading is less pronounced than in heterochromatin, it is nonetheless clearly detectable over 200 bp beyond siRNA-targeted sequences. Moreover, our analysis suggests that DNA methylation spreads from the center of euchromatic TE sequences towards their extremities, which are often not associated with siRNAs (Supplementary Figure S5) unlike their heterochromatic counterparts (Figure 6d). DISCUSSION Using a reﬁned version of our previous annotation pipeline, we have obtained the most extensive dataset for TE sequences in the Arabidopsis genome to date. Given the high sensitivity and speciﬁcity of this pipeline, this dataset is not expected to evolve substantially in the future. Similarly, the use of stringent criteria for the analysis of DNA methylation of TE sequences makes our conclusions particularly robust, and should facilitate comparison of DNA methylation patterns between different conditions as well as between Arabidopsis accessions. Furthermore, the methods used to determine the DNA methylation status of individual TE sequences based on bisulphite- sequencing data are general and can be implemented to the systematic analysis of the association between DNA methylation and any annotated features of genomes for which single-nucleotide resolution methylomes are avail- able (29). Based on the 17 085 TE sequences (out of a total of 31 245) for which DNA methylation could be examined with high precision, we have found that 26% are un- methylated and another 15% have methylation patterns that depart signiﬁcantly from the dense CG, CHG and CHH methylation typically reported. These two categories of TE sequences mainly correspond to short and highly degenerate relics located in euchromatin, many of which are missed by less sensitive detection pipelines. These relics