Figure 4

Identification of functionally important extracellular regions of carcinoembryonic antigen related cell adhesion molecules (CEACAMs) by multispecies sequence comparisons. N domain exon nucleotide sequences from orthologous CEACAMs of a selected set of mammalian species were aligned using the program ClustalW and the results were displayed as unrooted dendrogram (A). Note the high and low conservation of the CEACAM16 N2 and CEACAM1 N nucleotide sequences, respectively, as reflected by the size of the colored area. In order to determine the selective pressure on the maintenance of the nucleotide sequences, the number of nonsynonymous nucleotide substitution per nonsynonymous site (dN) and the number of synonymous substitutions per synonymous site (dS; B) as well as the accumulation of nonsynonymous and synonymous substitutions along immunoglobulin (Ig) variable- and constant-like domain exons (C) were determined after manual removal of sequence gaps for the following species: cattle, dog, human, mouse, opossum, rat and rhesus macaque (due to missing sequence information for the rhesus macaque CEACAM1 A2 exon, the corresponding exon from cat was used instead). The dN/dS ratios were calculated after manual editing of sequence gaps or insertions guided by the amino acid sequences for all branches of the resulting phylogenetic trees and displayed as mean values and standard deviations. Whole N domain exons were used for analysis including the regions (the first 12 codons) which encode part of the leader. Due to the variable truncation of the CEACAM20 N domain exons, the first common 27 codons were analysed. The mean dN/dS ratios were calculated and displayed as red (N domain exons) or blue columns (Ig constant-like domain exons); standard deviations are shown as bars. dN/dS values significantly greater than 1 (above the red line) indicate selective pressure for variability, a ratio less than 1 indicates pressures to conserve the protein sequence. Regions with high and low accumulation rates of nonsynonymous substitutions have been marked exemplarily in some graphs by black and red dotted lines. Note that the synonymous substitutions accumulate at the same rate for all exons.