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All the supplementary information are included in this file.

Supplementary methods, Figure S1-S8 and Table S1-S6

Ribonucleic acid export 1 (RAE1) plays an important role in the export of mature mRNAs from the nucleus to the cytoplasm. Long terminal repeats (LTRs) became integrated into the human genome during primate evolution. One such repeat element, LTR12C, lies within a predicted regulatory region located upstream of the RAE1 gene. We examined the transcriptional activity of LTR12C by using the luciferase assay, and showed that the tandem repeat region (TRR) located within LTR12C was required for its regulatory function. A bioinformatics analysis revealed that the LTR12C element had multiple transcription factor binding sites specific for nuclear transcription factor Y (NF-Y), and the promoter activity of LTR12C was significantly decreased after NF-Y knockdown. Additionally, we discovered novel data indicating that LTR12C was initially inserted into the gorilla genome. Taken together, our results reveal that the TRR of LTR12C has powerful regulatory activity due to its NF-Y binding sites, and the integration of the LTR12C element into the primate genome during evolution may have affected RAE1 transcription.

Table S1. A2 ratio in RH1 and LWS pigments estimated from MSP data. (PDF 119Â kb)

Figure S4. Amino acid alignment of LWS from river species. Residue positions are numbered according to the sequences of LWS. The dots and letters indicate identical and different residues, respectively, compared with the top line. The LWS sequences were determined from four riverine species: H. sp. ‘katonga’ (n = 2), H. sp. ‘katavi’ (n = 1), H. sp. ‘kitilda-rukwa’ (n = 2), and H. sp. ‘muzu-rukwa’ (n = 3). (PDF 22 kb)

Amino acid sequence variation in TAS2R38 of leaf-eating monkeys

Behavioral experiments, Genotyping of TAS2R38, and Expression vector and site-directed mutagenesis

Responses of colobines (N. larvatus B) with mutant TAS2R38 mimicking the macaque-type receptor.

Figure S1. Short descriptions of the ten species and the frequencies of LWS and RH1 alleles in the populations of (a) Haplochromis xenognathus, (b) H. sp. ‘green dentex’, (c) H. sp. ‘paropius like’, (d) Platytaeniodus degeni, (e) H. sp. ‘stone’, (f) H. piceatus, (g) H. sp. cf. hiatus, (h) H. sp. ‘supramacrops’, (i) H. sp. ‘deepwater cinctus’, and (j) H. sp. cf. fusiformis are shown in separate panels. Arabic numerals correspond to those in Fig. 1a, and the depths at each point are described on the right side of the numbers. The size of a pie indicates the number of haplotypes sequenced. The standard sizes of pies are shown at the left bottom. The colored sections of a pie indicate the frequency of the correspondent allele in the standard allele color pie (right bottom). The amino acid differences among allele groups are shown in Fig. 2b (LWS) and Fig. 2c (RH1). The maps were drawn by Y. T. based on original source maps: https://www.google.com/maps . Fish photographs were taken by M. I. and S. M. (PDF 3 kb)

Figure S2. Alignment of all polymorphic sites of LWS from the ten species. The nucleotide sites are shown on the top of the alignment. “n” and “s” indicate nonsynonymous and synonymous sites, respectively. Dots indicate the nucleotides that are identical with those in the top line. The allele groups of each sequence are shown on the right side of the sequences. (PDF 64 kb)

Figure S3. An alignment of all polymorphic sites of RH1 from 10 species used in this study. The nucleotide sites are shown on top of the alignment. “n” and “s” indicate nonsynonymous and synonymous sites, respectively. Dots indicate where nucleotides are identical with those in the top line. The allele groups of each sequence are shown right side of the sequences. (PDF 62 kb)

Diagnosis and remarks of species used in this study. (DOCX 21Â kb)

Sequence alignment of extant and ancestral TAS2R1 and TAS2R4 of New World monkeys (NWMs). (A) Sequence alignment of extant and ancestral NWM TAS2R1 receptors. (B) Sequence alignment of extant and ancestral NWM TAS2R4 receptors. Abbreviations of the species names for sequences are as follows: Cj, common marmoset (Callithrix jacchus); Cc, white-faced capuchin (Cebus capucinus); Aa, Azara's owl monkey (Aotus azarae); Ag, black-handed spider monkey (Ateles geoffroyi); Ap, mantled howler (Alouatta palliata). (DOCX 742 kb)

Reconstruction of ancestral TAS2R1s (A) and TAS2R4s (B). Ancestral amino acid sequences of New World Monkey TAS2R1 and TAS2R4 were inferred using the maximum likelihood method implemented in MEGA5 with the Dayhoff and JTT amino acid substitution models (Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S. Mol Biol Evol 2011; 28(10):2731-2739). (DOCX 237 kb)

Reconstruction of ancestral TAS2R1s (A) and TAS2R4s (B). Ancestral amino acid sequences of New World Monkey TAS2R1 and TAS2R4 were inferred using the maximum likelihood method implemented in MEGA5 with the Dayhoff and JTT amino acid substitution models (Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S. Mol Biol Evol 2011; 28(10):2731-2739). (DOCX 237 kb)