Automated conserved non-coding sequence (CNS) discovery reveals differences in gene content and promoter evolution among grasses

Abstract

Conserved non-coding sequences (CNS) are islands of non-coding sequence that, like protein coding exons, show less divergence in sequence between related species than functionless DNA. Several CNSs have been demonstrated experimentally to function as cis-regulatory regions. However, the specific functions of most CNSs remain unknown. Previous searches for CNS in plants have either anchored on exons and only identified nearby sequences or required years of painstaking manual annotation. Here we present an open source tool that can accurately identify CNSs between any two related species with sequenced genomes, including both those immediately adjacent to exons and distal sequences separated by >12 kb of non-coding sequence. We have used this tool to characterize new motifs, associate CNSs with additional functions, and identify previously undetected genes encoding RNA and protein in the genomes of five grass species. We provide a list of 15,363 orthologous CNSs conserved across all grasses tested. We were also able to identify regulatory sequences present in the common ancestor of grasses that have been lost in one or more extant grass lineages. Lists of orthologous gene pairs and associated CNSs are provided for reference inbred lines of arabidopsis, Japonica rice, foxtail millet, sorghum, brachypodium, and maize.

RNA-directed DNA methylation enforces boundaries between heterochromatin and euchromatin in the maize genome

“…We hypothesized that similar boundaries may also be required at regulatory regions to allow access to these regions for transcription factors. Turco et al (17) identified a number of CNSs in the B73 genome and 11,680 of these are located >5 kb from the nearest gene. The profile of DNA methylation relative to these CNSs >5 kb away from genes reveals the presence of mCHH islands flanking CNSs and reduced DNA methylation at CNSs (Fig.…”

Differentially Regulated Orthologs in Sorghum and the Subgenomes of Maize

“…As transcription factors are often associated with larger quantities of conserved noncoding sequences (CNSs) (Freeling et al, 2007;Turco et al, 2013), we also investigated the number and quantity of conserved noncoding sequence associated with different classes of genes; however, no strong patterns were observed ( Figure 6D). The use of conserved noncoding sequence data to identify regulatory sequence requires that the regulatory sequence be conserved between species.…”

“…CNSs were identified using the CNS Discovery Pipeline 3.0 (CDP) (Turco et al, 2013) with some modifications. Specifically, the built-in syntenic gene identification pipeline from the CDP was replaced with the previously defined syntenic gene list described above.…”

The Most Deeply Conserved Noncoding Sequences in Plants Serve Similar Functions to Those in Vertebrates Despite Large Differences in Evolutionary Rates

“…In the second approach to detecting deeply conserved CNSs, peach-cacao orthologous CNSs identified using the PL3.0 automated CNS Discovery pipeline (Turco et al, 2013) were used to search for BLASTN hits present in the 10,000-bp region surrounding columbine and Arabidopsis orthologous genes. This approach allowed CNSs to be identified from Arabidopsis genes in which one Approximate points of divergence and WGD events (ellipses) are shown in million years (Myr), with a filled-in ellipse indicating the paleohexaploidy event that occurred in the ancestor of most eudicots.…”