Fellowships Grants and Awards May 2004

Background: The recent availability of genome sequences has provided unparalleled insights into the broad-scale patterns of transposable element (TE) sequences in eukaryotic genomes. Nevertheless, the difficulties that TEs pose for genome assembly and annotation have prevented detailed, quantitative inferences about the contribution of TEs to genomes sequences. Results: Using a high-resolution annotation of TEs in Release 4 genome sequence, we revise estimates of TE abundance in Drosophila melanogaster. We show that TEs are non-randomly distributed within regions of high and low TE abundance, and that pericentromeric regions with high TE abundance are mosaics of distinct regions of extreme and normal TE density. Comparative analysis revealed that this punctate pattern evolves jointly by transposition and duplication, but not by inversion of TE-rich regions from unsequenced heterochromatin. Analysis of genome-wide patterns of TE nesting revealed a 'nesting network' that includes virtually all of the known TE families in the genome. Numerous directed cycles exist among TE families in the nesting network, implying concurrent or overlapping periods of transpositional activity. Conclusion: Rapid restructuring of the genomic landscape by transposition and duplication has recently added hundreds of kilobases of TE sequence to pericentromeric regions in D. melanogaster. These events create ragged transitions between unique and repetitive sequences in the zone between euchromatic and beta-heterochromatic regions. Complex relationships of TE nesting in beta-heterochromatic regions raise the possibility of a co-suppression network that may act as a global surveillance system against the majority of TE families in D. melanogaster. Background Nearly all eukaryotic genomes contain a substantial fraction of middle repetitive, transposable element (TE) sequences interspersed with the unique sequences encoding genes and cis-regulatory elements. The broad-scale patterns of TE abundance and distribution in various model organisms have Published: 29 November 2006 Genome Biology 2006, 7:R112 (doi:10.1186/gb-2006-7-11-r112) Received: 31 July 2006 Revised: 13 November 2006 Accepted: 29 November 2006 The electronic version of this article is the complete one and can be found online at http://genomebiology.com/2006/7/11/R112 Genome Biology 2006, 7:R112 R112.2 Genome Biology 2006, Volume 7, Issue 11, Article R112 Bergman et al. http://genomebiology.com/2006/7/11/R112 become increasingly well-understood with the recent availability of essentially complete genome sequences (for example, [1-4]). Despite these general advances, however, a detailed understanding of the evolutionary forces that control the abundance and distribution of TEs remains elusive, owing in part to the dynamic nature of this component of the genome as well as to the inherent problems that TE sequences present for genome assembly and annotation. As with all unfinished whole-genome shotgun assemblies, uncertainty in the assembly of repetitive DNA in the first two releases of the Drosophila melanogaster genome sequence posed difficulties for analysis of TE sequences [5-8]. The improved assembly of repetitive regions in the D. melanogaster Release 3 genome sequence presented the first opportunity to study TEs in a finished whole genome shotgun sequence [2,9], revealing the true challenge that these sequences pose for their systematic annotation [10,11]. With further improvements in the Release 4 genome sequence made possible by the efforts of the Berkeley Drosophila Genome Project [12] (especially in regions of high TE density where several gaps have been completed), we are now in a position to establish more stable trends in TE abundance for D. melanogaster. In addition to having access to improved genome sequence data, we have recently developed an improved TE annotation pipeline that uses the combined evidence of multiple computational methods to predict 'TE models' in genome sequences [10]. We have shown that this pipeline identifies a large number of predicted TEs that were omitted from the Release 3 genome annotations, and subsequently applied this system to the D. melanogaster Release 4 sequence [10]. Here we analyze the results of this effort in detail, which allows an extremely high-resolution view of the structure and location of TEs in one of the highest quality metazoan genome sequences currently available. We first revised baseline estimates of the TE abundance in the Drosophila genome sequence, based on the fact that TEs show a strikingly non-random distribution across the genome. We then used this baseline to identify specific regions of extremely high TE density in the genome sequence. This analysis showed that regions of the genome broadly known to have high TE abundance, such as pericentromeric regions and the fourth chromosome, are in fact often characterized by distinctly localized regions of extremely high TE density interrupted by regions of lower TE density. Comparative sequence analysis showed that this punctate pattern is unlikely to have arisen in the D. melanogaster genome by inversion of TE-rich heterochromatic sequences, but can evolve in situ by the joint action of recurrent transposition and duplication. Finally, we analyzed in detail the patterns of TE nesting in the genome sequence, taking advantage of the improved joining of fragments from the same TE insertion event in our new annotation. We framed the process of TE nesting as a directed graph and borrowed techniques from network analysis to study genome-wide patterns of TE nesting. This work demonstrates the added value of high-resolution annotations for understanding how TEs impact genome organization and evolution, and preludes the interpretation of TE-rich heterochromatic regions currently being sequenced by the Drosophila Heterochromatin Genome Project [13]. Results Abundance and distribution of TEs in the Release 4 genome sequence Using a recently completed combined-evidence annotation of the Release 4 genome sequence [10], we revised estimates of the overall abundance of TE sequences in D. melanogaster (Table 1) from those based on the Release 3 sequence [2]. Excluding foreign elements based on query sequences from other species (see Materials and methods), the estimated number of TEs in the D. melanogaster Release 4 genome sequence (n = 5,390) is over three-fold higher than in Release 3 (n = 1,572). In contrast, the amount of sequence annotated as TE increased by only approximately 44% in Release 4 (6.51 Mb, 5.50% of genome) relative to Release 3 (4.51 Mb, 3.86% of genome). (We note that the proportion of the Release 4 genome estimated here as TE is calculated as the sum of nonredundant annotation spans including unique sequences inserted into TEs; this procedure differs slightly from our previous estimates for Release 4, which only included sequences strictly homologous to TE query sequences [10].) The discrepant changes in these two metrics of TE abundance across releases results from the fact that almost all new TEs in Release 4 are either small fragments and/or annotations of the highly abundant but degenerated INE-1 element (also known as DINE-1 or DNAREP1_DM) [14], a family that was omitted from the Release 3 annotation. The inclusion of these new small fragments is also reflected in the fact that the proportion of TEs estimated to be full-length (defined as ± 3% of the canonical element including the length of inserted sequences) has declined from 30.5% in Release 3 to 9.83% in Release 4. The number of TEs involved in nests (n = 785) has more than doubled in Release 4 relative to Release 3 because of newly annotated sequences and improved joining of TE fragments belonging to the same insertion, although the estimated proportion of TEs involved in nests (14.6%) in Release 4 has decreased relative to Release 3 as a consequence of the increased total number of TEs annotated. The major patterns of TE abundance identified in previous releases of the D. melanogaster genome sequence [2,7,8,15,16] are also observed in Release 4, suggesting that these trends are stable features of the D. melanogaster genomic landscape. As shown in Figure 1, both the pericentromeric regions of the major chromosome arms and the entirety of chromosome 4 have higher densities of TE insertions, relative to non-pericentromeric regions [2,7,15]. Densities over the non-pericentromeric regions are roughly equal, with no general increase in TE density in telomeric regions Genome Biology 2006, 7:R112 http://genomebiology.com/2006/7/11/R112 Genome Biology 2006, Volume 7, Issue 11, Article R112 Bergman et al. R112.3