Analysis of conserved noncoding DNA in Drosophila reveals similar constraints in intergenic and intronic sequences

Comparative genomic approaches to gene and cis-regulatory prediction are ba sed on the principle that differential DNA sequence conservation reflects v ariation in functional constraint. Using this principle, we analyze noncodi ng sequence conservation in Drosophila for 40 loci with known or suspected cis-regulatory function encompassing > 100 kb of DNA. We estimate the fract ion of noncoding DNA conserved in both intergenic and intronic regions and describe the length distribution of ungapped conserved noncoding blocks. Oi l average, 22%-26% of noncoding sequences surveyed are conserved in Drosoph ila, with median block length similar to 19 bp. We show that point Substitu tion in conserved noncoding blocks exhibits transition bias as well as line age effects in base composition, and Occurs more than all order of magnitud e more frequently than insertion/deletion (indel) substitution. Overall, pa tterns of noncoding DNA structure and evolution differ remarkably little be tween intergenic and intronic conserved blocks, suggesting that the effects of transcription per se contribute minimally to the constraints operating on these Sequences. The results of this study have implications for the dev elopment of alignment and prediction algorithms specific to noncoding DNA, as well as for models of cis-regulatory DNA sequence evolution.