Comparison of Ultra-Conserved Elements in Drosophilids and Vertebrates

Igor V Makunin,Stepan N Belyakin,Stuart J Stephen,Viktor V Shloma,Michael Pheasant,Denis Dupuy

doi:10.1371/journal.pone.0082362

Abstract

Metazoan genomes contain many ultra-conserved elements (UCEs), long sequences identical between distant species. In this study we identified UCEs in drosophilid and vertebrate species with a similar level of phylogenetic divergence measured at protein-coding regions, and demonstrated that both the length and number of UCEs are larger in vertebrates. The proportion of non-exonic UCEs declines in distant drosophilids whilst an opposite trend was observed in vertebrates. We generated a set of 2,126 Sophophora UCEs by merging elements identified in several drosophila species and compared these to the eutherian UCEs identified in placental mammals. In contrast to vertebrates, the Sophophora UCEs are depleted around transcription start sites. Analysis of 52,954 P-element, piggyBac and Minos insertions in the D. melanogaster genome revealed depletion of the P-element and piggyBac insertions in and around the Sophophora UCEs. We examined eleven fly strains with transposon insertions into the intergenic UCEs and identified associated phenotypes in five strains. Four insertions behave as recessive lethals, and in one case we observed a suppression of the marker gene within the transgene, presumably by silenced chromatin around the integration site. To confirm the lethality is caused by integration of transposons we performed a phenotype rescue experiment for two stocks and demonstrated that the excision of the transposons from the intergenic UCEs restores viability. Sequencing of DNA after the transposon excision in one fly strain with the restored viability revealed a 47 bp insertion at the original transposon integration site suggesting that the nature of the mutation is important for the appearance of the phenotype. Our results suggest that the UCEs in flies and vertebrates have both common and distinct features, and demonstrate that a significant proportion of intergenic drosophila UCEs are sensitive to disruption.

Highlights

Comparative analysis of mammalian and insect genomes have demonstrated that the majority of the evolutionarily constrained sequences in these lineages are located outside of protein coding regions [1,2]
In our work we focus on the comparison of Ultra-Conserved Elements (UCEs) in sets of species with similar phylogenetic distances
We compared the UCEs in vertebrates and drosophilids and found that both the number and length of UCEs are smaller in insects, in agreement with previous reports [44,45]

Summary

Introduction

Comparative analysis of mammalian and insect genomes have demonstrated that the majority of the evolutionarily constrained sequences in these lineages are located outside of protein coding regions [1,2]. Comparison of the human and mouse sequences on chromosome 21 showed that many non-coding sequences are even more conserved than protein-coding regions [3]. Subsequent studies identified numerous highly conserved non-coding elements (CNEs) in species as evolutionarily distant as human and fish [4,5], which are clustered around genes involved in regulation of transcription and development [6]. The Ultra-Conserved Elements (UCEs) are arguably the most constrained sequences in the human genome. The UCEs were first identified as sequences at least 200 bp long identical between the human, mouse and rat genomes [7]. An alignmentindependent method was used for the identification of both syntenic and non-syntenic Long Identical Multispecies Elements in vertebrates and plants [10] including some elements omitted in earlier studies due to gap alignment deficiencies

Methods

Results

Conclusion