Molecular evolution of Drosophila Cdc6, an essential DNA replication-licensing gene, suggests an adaptive choice of replication origins

Abstract

Increased size of eukaryotic genomes necessitated the use of multiple origins of DNA replication, and presumably selected for their efficient spacing to ensure rapid DNA replication. The sequence of these origins remains undetermined in metazoan genomes, leaving important questions about the selective constraints acting on replication origins unanswered. We have chosen to study the evolution of proteins that recognize and define these origins every cell cycle, as a surrogate to the direct analysis of replication origins. Among these DNA replication proteins is the essential Cdc6 protein, which acts to license origins for replication. We find that two different species pairs of Drosophila show evidence of positive selection in Cdc6 in their highly conserved C-terminal AAA-ATPase domain. We also identified amino acid segments that are highly conserved in the N-terminal tail of Cdc6 proteins from various Drosophila species, but are not conserved even in closely related insect species. Instead, we find that the N-terminal tails of Cdc6 proteins vary extensively in size and sequence across different eukaryotic lineages. Our results suggest that choice of origin firing may be significantly altered in closely related species, as each set of replication proteins optimizes to its own genomic landscape.

Keywords: Drosophila melanogaster; Drosophila pseudoobscura; McDonald-Kreitman test; Saccharomyces cerevisiae; non-coding DNA; origin ‘choice’; replication origins.

Figure 1. Cdc6 phylogeny

A neighbor-joining tree based on an amino acid alignment of the Cdc6 proteins in Drosophila is presented. Bootstrap support (percentage of 1,000 trials) is shown next to each node. This phylogeny is largely congruent with that of Drosophila species.

Figure 2. Evolution of Cdc6’s N-terminal tail

(A) Schematic representation of Cdc6 from different Drosophila species. The N-terminal tail is shown as a series of conserved sequence blocks (numbered 1 through 5) and a thin line, while the C-terminal domain (consisting of the AAA+ ATPase and winged helix-like DNA binding domains) is shown in a gray box. The position of the intron conserved through all the Drosophila species is shown with a thin vertical line. (B) The 5 conserved blocks schematized in B are presented in Logos format. Note that position 11 in Block 3 is a gapped position.