The genomic organization and deduced sequence of hTRPC2. (A) Structure of the TRPC2 gene. Thirteen exons were identified on chromosome 11p15.5 in a draft sequence of the human genome. Black boxes represent exons containing coding sequence. White boxes indicate exons that contain noncoding sequence; the size of these exons was not determined. Breaks in the sequence represent regions of the human genome for which the data are still ambiguous. The positions of primer pairs I–V are indicated. Note that two primer pairs (III and IV) span introns. (B) The deduced sequence of hTRPC2 aligned with the sequence of mouse TRPC2 (mTRPC2). Identical residues are boxed in black. The position of intron/exon boundaries is indicated below the sequence. Vertical arrows indicate the positions of deleterious mutations. Site 1 is a 4-bp insertion and site 2 is a 1-bp deletion. These nucleotides were removed to produce the coding sequence shown. Stop codons are indicated by an asterisk. Bars underlining sequences indicate the portion of the sequence used for determining K_a/K_s.

Inferred time of occurrence of deleterious mutations in TRPC2 during primate phylogeny. (A) Mutations are numbered 1–9. Mutations 1–6 are found in human at the positions indicated in Fig. 1. Mutation 6 was found in all OW monkeys and hominoids but not in NW monkeys or prosimians; therefore, we infer that it occurred in the common ancestor of OW monkeys and apes. Mutation 9 is a missense mutation that was found in all OW monkeys and in gibbon but not in other hominoids, suggesting either that a reversion event occurred more recently in evolution (indicated by a black circle with a white 9 on the tree) or that the mutation arose independently more than once. Two frameshift mutations (7 and 8) were found only in OW monkeys. Mutation 7 is a 1-bp insertion at a position equivalent to mouse D483 and mutation 8 is a 13-bp deletion spanning the same position. The deletion event therefore restores the reading frame. (B) A schematic representation of the TRPC2 ion channel indicating the position of each mutation. Black bars represent the transmembrane domains.

Rates of synonymous and nonsynonomous substitutions in the TRPC2 gene across primate phylogeny. Pairwise comparisons of sequence from all species were performed and K_a/K_s values were computed. Within-group values show the average K_a/K_s ratio for all pairwise comparisons in a group. A double asterisk indicates that K_a/K_s is not compatible with neutral evolution (P ≤ 0.002). The K_a/K_s values computed by using predicted ancestral sequences are shown above the corresponding branch of the tree, and the total number of substitutions is shown in parentheses. Branches on which selective pressure was strongly evident (P ≤ 0.002) are shown with a thick line, and K_a/K_s values are marked with a double asterisk. This P value was derived by using the Bonferroni correction to ensure an overall type I error rate (false rejection of the null hypothesis) of 5% for each of the 22 tests conducted. On two branches, K_a/K_s values were significantly different from unity at P < 0.05. These branches are marked with a single asterisk. Lengths of branches are proportional to divergence dates for lineages of NW monkeys and OW monkeys and apes. Dates are based on ref. 34. Ro, rodent; Pr, prosimian; mya, million years ago.