Abstract
BACKGROUND: Mouse E11.5 embryonic heart enhancers were found to exhibit exceptionally weak sequence conservation during vertebrate evolution compared to enhancers of other developing organs. However, it is unknown whether this phenomenon is due to elevated mutation rates, or is a consequence of natural selection. RESULTS: In this study, based on the aligned orthologous genomic sequences of mouse and other closely related mammals, the substitution rates of fourfold degenerate sites or intron sequences in neighboring genes were used as neutral references to normalize substitution rates of mouse enhancers. Subsequent comparisons indicated that heart enhancers' evolutionary rates were increased by natural selection. Correspondingly, the results of Fisher's exact tests to examine the differential enrichment of substitutions between enhancers and neutral sequences suggest that both relaxed purifying selection and positive selection caused the rapid evolution of heart enhancers. Analyses on recombination rates and substitution patterns indicated that GC-biased gene conversion does not contribute to evolutionary rate variations among enhancers. In general, pleiotropic enhancers and enhancers in proximity to weakly expressed genes, tend to evolve slowly. Although heart enhancers are less pleiotropic and are adjacent to highly expressed genes, these biases do not account for the rapid evolution observed. CONCLUSIONS: In combination, the results of the present study suggest that factors associated with functions or characteristics of the tissue may exert direct and profound effects on the intensity and direction of the natural selection applied to regulatory DNAs, such as enhancers.