Computational methods to detect conserved non-genic elements in phylogenetically isolated genomes: application to zebrafish

Abstract

Many important model organisms for biomedical and evolutionary research have sequenced genomes, but occupy a phylogenetically isolated position, evolutionarily distant from other sequenced genomes. This phylogenetic isolation is exemplified for zebrafish, a vertebrate model for cis-regulation, development and human disease, whose evolutionary distance to all other currently sequenced fish exceeds the distance between human and chicken. Such large distances make it difficult to align genomes and use them for comparative analysis beyond gene-focused questions. In particular, detecting conserved non-genic elements (CNEs) as promising cis-regulatory elements with biological importance is challenging. Here, we develop a general comparative genomics framework to align isolated genomes and to comprehensively detect CNEs. Our approach integrates highly sensitive and quality-controlled local alignments and uses alignment transitivity and ancestral reconstruction to bridge large evolutionary distances. We apply our framework to zebrafish and demonstrate substantially improved CNE detection and quality compared with previous sets. Our zebrafish CNE set comprises 54 533 CNEs, of which 11 792 (22%) are conserved to human or mouse. Our zebrafish CNEs (http://zebrafish.stanford.edu) are highly enriched in known enhancers and extend existing experimental (ChIP-Seq) sets. The same framework can now be applied to the isolated genomes of frog, amphioxus, Caenorhabditis elegans and many others.

Robust Identification of Developmentally Active Endothelial Enhancers in Zebrafish Using FANS-Assisted ATAC-Seq

“…1H). These zCNEs were identified through local alignments of multiple fish and tetrapod genomes to detect remote homology between zebrafish and at least two other species (Hiller et al, 2013). In all cases, concordance between ATAC-Seq reads and histone marks or zCNEs were obtained for the replicate libraries (Fig.…”

“…Previous studies have demonstrated the utility of using conservation to identify endothelial enhancers in several zebrafish studies (Bussmann et al, 2010; Veldman and Lin, 2012). However, the lack of closely related genomes for accurate homology alignment limits a more comprehensive identification of putative enhancers using this approach (Hiller et al, 2013). Indeed, analysis of our endothelial ATAC-Seq elements indicate that more than 80% do not bear known conserved sequences, suggesting that available CNE annotations only detect a very small proportion of tissue-specific enhancer elements.…”

“…Subsequent availability of fish genome sequences, along with phylogenetic footprinting (Wasserman et al, 2000) enabled identification of small conserved non-coding elements (CNEs), which often possessed enhancer activity (Gottgens et al, 2002; Komisarczuk et al, 2009). However, unlike Drosophila and many mammalian species, for which closely related genomes are available for accurate comparison, the zebrafish is considered to be phylogenetically isolated regarding available genome sequences (Hiller et al, 2013). As a consequence, annotation of CNEs in the zebrafish genome is less comprehensive compared to other model systems.…”

Differential DNA methylation at conserved non-genic elements and evidence for transgenerational inheritance following developmental exposure to mono(2-ethylhexyl) phthalate and 5-azacytidine in zebrafish

“…We used a computationally derived list of zfCNEs which contained over 54,000 regions [35]. CNEs are generally located outside genic regions and can have cis-regulatory functions such as enhancers or silencers [36].…”

A Comprehensive Review of Emerging Computational Methods for Gene Identification

“…Comparative methods [166], including pairwise and multiple-wise alignment among ancestrally constructed species, have been adopted to detect non-genic elements. By detecting thousands of conserved non-genic elements and revealing the majority of evolutionarily conserved DNA sequences, a computational framework based on alignments has been applied to zebra-fish, a phylogenetically isolated genome.…”