Regulatory conservation of protein coding and microRNA genes in vertebrates: lessons from the opossum genome

Abstract

Background: Being the first noneutherian mammal sequenced, Monodelphis domestica (opossum) offers great potential for enhancing our understanding of the evolutionary processes that take place in mammals. This study focuses on the evolutionary relationships between conservation of noncoding sequences, cis-regulatory elements, and biologic functions of regulated genes in opossum and eight vertebrate species.

Results: Analysis of 145 intergenic microRNA and all protein coding genes revealed that the upstream sequences of the former are up to twice as conserved as the latter among mammals, except in the first 500 base pairs, where the conservation is similar. Comparison of promoter conservation in 513 protein coding genes and related transcription factor binding sites (TFBSs) showed that 41% of the known human TFBSs are located in the 6.7% of promoter regions that are conserved between human and opossum. Some core biologic processes exhibited significantly fewer conserved TFBSs in human-opossum comparisons, suggesting greater functional divergence. A new measure of efficiency in multigenome phylogenetic footprinting (base regulatory potential rate [BRPR]) shows that including human-opossum conservation increases specificity in finding human TFBSs.

Conclusion: Opossum facilitates better estimation of promoter conservation and TFBS turnover among mammals. The fact that substantial TFBS numbers are located in a small proportion of the human-opossum conserved sequences emphasizes the importance of marsupial genomes for phylogenetic footprinting-based motif discovery strategies. The BRPR measure is expected to help select genome combinations for optimal performance of these algorithms. Finally, although the etiology of the microRNA upstream increased conservation remains unknown, it is expected to have strong implications for our understanding of regulation of their expression.

Figure 1

Phylogenetic tree of the species examined in this study. This phylogenetic tree is based on the University of California, Santa Cruz (UCSC) multiple alignments. The tree was generated using phyloGif [72].

Figure 2

Upstream sequence conservation of protein coding versus miRNA genes. Comparison of 5-kilobase upstream sequence conservation between human and various organisms, relative to the transcription start site (TSS; protein-coding, solid blue line) and gene start (intergenic microRNA [miRNA] genes, orange line). The conservation of developmental genes (light blue dotted line) and tRNA genes (green dotted line) are also plotted for comparison purposes. For the plot 100 base pair (bp) intervals were used for the first 500 bp and 500 bp intervals thereafter.

Figure 3

Conserved block coverage of the 5 kilobases upstream regions versus TFBS turnover rates. A third-order polynomial trendline is fitted for illustration. TFBS, transcription factor binding site.

Figure 4

Association between BRPR scores and detectable sites. For each given percent of detectable transcription factor binding sites (TFBSs), the combination of aligned genomes with the highest base regulatory potential rate (BRPR) value will yield the smaller conserved region (for phylogenetic footprinting algorithm searches). The full list of genome combinations and their BRPR values are given in Additional data file 1. The blue line presents the association between percentage of human TFBSs located in conserved regions in a combination of genomes with this BRPR value among all possible genome combinations in this study (see text for detailed description). The grey line plot is similar after the opossum genome is omitted (see text). BRPR, base regulatory potential rate.

Figure 5

Cross-species conservation of individual TFBS positions versus their information content. Conservation is measured between the human and each of the other species. Information content is measured according to the human position-specific score matrix (PSSM) model.