Visual Pigment Genes: Evolution - Wiley Online Library

Visual Pigment Genes: Evolution

Visual Pigment Genes: Evolution Shozo Yokoyama, Syracuse University, Syracuse, New York, USA

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More than 100 visual pigment genes have been cloned from a diverse range of vertebrates. Comparative sequence analyses of these genes and in vitro assays of engineered visual pigments have been used to elucidate not only the molecular bases of color vision but also the processes of adaptive evolution at the molecular level.

Article contents Visual Pigment Genes and Visual Pigments Coelacanths and RH1 and RH2 Pigments LWS and MWS Pigments SWS1 Pigments Perspectives

Visual Pigment Genes and Visual Pigments Vision begins when photons are absorbed by photosensitive molecules, visual pigments. The visual pigments in rod cells are referred to as rhodopsin, whereas those in cone cells are often called cone pigments. Each visual pigment consists of an apoprotein, opsin and the chromophore, usually 11-cisretinal, whose spectral sensitivity is characterized by the wavelength of maximal absorption (lmax). The two molecules are bound to each other by a Schiff base linkage to the rhodopsin lysine residue, K296, or equivalent lysine of the cone pigments (Palczewski et al., 2000). The Schiff base of 11-cis-retinal is usually protonated by the glutamate counterion, E113, of the opsin. The protonated Schiff base has a lmax of 440 nm in solution. Interacting with an opsin, however, the Schiff base-linked chromophore in a visual pigment can have a lmax from 360 to 600 nm. This phenomenon is known as the spectral tuning of visual pigments. The opsin is encoded by a specific visual pigment gene (or an opsin gene). The cone pigment genes were first isolated from humans (Nathans et al., 1986). Using the corresponding opsin and rhodopsin complementary deoxyribonucleic acids (cDNAs), more than 100 complete opsin genes and cDNA clones have been isolated and sequenced. Based on their nucleotide and deduced amino acid sequences, the opsin genes (and visual pigments) in vertebrate retinas are classified into five evolutionary groups: (1) RH1 (rhodopsins); (2) RH2 (RH1-like); (3) short wavelength-sensitive type 1 (SWS1); (4) SWS type 2 (SWS2); and (5) long wavelength- and middle wavelength-sensitive (LWS/MWS) groups. The RH1 genes are usually expressed in rods and the other four groups of opsin genes usually in cones. The gene size ranges from approximately 1 kb of the fish RH1 genes to approximately 20 kb of the rat MWS gene (Figure 1). In the early stage of fish evolution, all introns of the RH1 genes were lost. Otherwise, the introns 1, 2, 3 and 4 of the RH1, RH2, SWS1 and SWS2 genes and introns 2, 3, 4 and 5 of the LWS/MWS genes interrupt

doi: 10.1002/ 97 8 0470 015902 . a 0006148

their coding sequences at exactly the same corresponding sites. The RH1 genes (and pigments) are most closely related to RH2, and then to the SWS2, SWS1 and LWS/MWS groups, in that order, which are strongly supported by high bootstrap values (Figure 2). In Figure 2, however, the phylogenetic positions of chameleon (P491) pigment in the RH1 group and gecko (P521) pigment in the LWS/MWS group do not agree with the phylogenetic relationships of organisms. The diurnal chameleon (Anolis carolinensis) has only cones, whereas the nocturnal gecko (Gekko gekko) has only rods. Thus, the incorrect phylogenetic positions of the two pigments seem to reflect their rapid mutant substitutions associated with the switch in the photoreceptor cell-specificity. Here two additional comments are in order. First, the five groups of genes have arisen through four gene-duplication events. The RH1 group contains visual pigments from a wide variety of organisms, ranging from lampreys to mammals. As the most recent gene duplication event of the four occurred prior to the divergence of various vertebrates, the vertebrate ancestor must have possessed all five groups of opsin genes. Second, the ability of humans to see light ranging in wavelength from 400 to 650 nm is controlled by the RH1 (human; P497), SWS1 (human; P414), MWS (human; P530) and LWS (human; P560) pigments (Figure 2). So far, neither RH2 nor SWS2 genes has been found in the human and other mammalian genomes. These genes must have become nonfunctional and been lost in an early stage of mammalian evolution. From Figure 2, we can also see that RH1, RH2, SWS1, SWS2 and LWS/MWS pigments have a lmax of 480–510, 470–510, 360–430, 410–460 and 510–560 nm. In the following discussion, the amino acid site numbers are those of the bovine (P500) pigment in the RH1 group.

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Visual Pigment Genes: Evolution RH1 Marine lamprey (P500) Cavefish (P503) Af. coelacanth (P485) Chameleon (P491) Chicken (P503) Pigeon (P502) Human (P497) Bovine (P500) Mouse (P498) RH2 Malawi fish-Dc (P536) Malawi fish-Mz (P533) Af. coelacanth (P478) Chameleon (P495) Pigeon (P503) SWS1 Malawi fish-Mz (P368) Chameleon (P358) Pigeon (P393) Human (P414) Squirrel monkey (P433) SWS2 Malawi fish-2A-Dc (P447) Malawi fish-2B-Dc (P447) Chameleon (P437)

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2 kb

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Pigeon (P448) LWS/MWS Cavefish (P563) Cavefish (P533) Malawi fish-Dc (P536) Chameleon (P561) Pigeon (P558) Human (P530) Human (P560) Human (P560) Goat (P553) Rabbit (P509) Rat (P509) Marmoset (P561) Marmoset (P553) Marmoset (P539)

Figure 1 Structures of visual pigment genes, where exons and introns are represented by black boxes and horizontal lines respectively. The numbers after P refer to lmax. For Malawi fish pigments, Dc and Mz denote Dimidiochromis compressiceps and Metriaclima zebra respectively. Malawi fish-Dc (P536), Malawi fish-Mz (P533), Malawi fish-2A-Dc (P447), Malawi fish-2B-Dc (P488), Malawi fish-Dc (P368), Malawi fish-Dc (P569), marmoset (P561), marmoset (P553) and marmoset (P539) pigment genes are from GenBank (accession nos. AF247121, AF247122, AF247113, AF247118, AF191220, AF247125, AB046549s1–s6, AB046555s1–s6 and AB046561s1–s6 respectively). For other genes, see Yokoyama (2000). The gene duplication of the human P530 and P560 genes occurred some 30 million years (MY) ago (Nathans et al., 1986). Two human (P560) genes have intron 1 length polymorphism, one of them being 2 kb longer than the other. Af: African; LWS/MWS: long wavelength- and middle wavelength-sensitive; RH1: rhodopsins; RH2: RH1-like; SWS1: short wavelengthsensitive type 1; SWS2: SWS type 2.

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RH1

RH2

SWS2

Malawi fish-Dc (P368) Zebrafish (P362) Goldfish (P359) Clawed frog (P425) Chameleon (P358) Pigeon (P393) Chicken (P415) Budgerigar (P363) Zebra finch (P359) Blackbird (P360) Canary (P366) Human (P414) Macaque (P431) Squirrel monkey (P430) Marmoset (P423) Bovine (P431) Mouse (P359) Rat (P358)

Bovine (P500) Dog (P508) Rat (P498) Mouse (P498) Rabbit (P502) Macaque (P500) Human (P497) Saddl dolphin (P489) Bnose dolphin (P488) Zebra finch (P502) Chicken (P503) Pigeon (P502) Alligator (P499) Bull frog (P500) Leopard frog (P502) Clawed frog (P502) Salamander (P506) Ind. coelacanth (P485) Af. ceolacanth (P485) John Dory (P492) Mosquitofish (P505) Guppy (P503) Sandgoby (P501) Zebrafish (P500) Carp (P499) Goldfish (P492) Cave fish (P503) Conger eel (P487) Marine eel (P482) River eel (P502) Skate (P500) Chameleon (P491) River lamprey (P500) Marine lamprey (P500) Malawi fish-Dc (P536) Malawi fish-Mz (P533) Zebrafish G1 (P480) Goldfish (P511) Goldfish (P506) Zebrafish G2 (P480) Af. coelacanth (P478) Ind. coelacanth (P478) Chameleon (P495) Pigeon (P503) Chicken (P508) Zebra finch (P505) Gecko (P467) Malawi fish-2A-Dc (P447) Malawi fish-2B-Dc (P447) Cave fish (P432) Goldfish (P441) Zebrafish (P415) Bull frog (P432) Salamander (P431) Chameleon (P437) Pigeon (P448) Chicken (P455) Zebra finch (P441)

0.1 : Bootstrap > 0.9

SWS1

Gecko (P521) Cave fish G101 (P533) Cave fish G103 (P533) Malawi fish-Dc (P536) Cave fish (P563) Goldfish (P559) Zebrafish (P570) Clawed frog (P557) Chameleon (P561) Pigeon (P558) Chicken (P561) Zebra finch (P560) Human (P530) Human (P560) Capuchin (P535) Capuchin (P550) Capuchin (P561) Squirrel monkey (P561) Marmoset (P556) Marmoset (P561) Tamarin (P561) Tamarin (P556) Marmoset (P543) Tamarin (P543) Squirrel monkey (P535) Squirrel monkey (P550) Goat (P553) Deer (P531) Bottlenose dolphin (P524) Cat (P553) Horse (P545) Rabbit (P509) Guinea pig (P516) Squirrel (P532) Mouse (P508) Rat (P509) Mole rat (P534)

LWS/MWS

Figure 2 Phylogenetic tree for the vertebrate visual pigments by applying the neighbor-joining method (Saitou and Nei, 1987) to their amino acid sequences. Ind. coelacanth (P485) and Ind. coelacanth (P478) are from Indonesian coelacanth (Latimeria menadoensis. Salamander (P431), bull frog (P432) and mole rat (P534) pigments are from GenBank (accession nos. AF038946, AB010085 and AF139726 respectively). Blackbird (P360) is from red-winged blackbird (Agelaius pheniceus). For other sequences, see Yokoyama (2000). The arrow indicates the root of the phylogenetic tree. The bar at the bottom indicates evolutionary distance measured as the number of amino acid replacements per site.

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Coelacanths and RH1 and RH2 Pigments How did organisms modify their color vision to adapt to various environments? This evolutionary question is closely related to a central question in phototransduction: How do visual pigments detect a wide range of wavelengths using the same 11-cis-retinal? Thus, evolutionary biology and vision science have an important common goal. Shortly after the cloning of the human cone pigment genes, the functional assay of visual pigments was developed, where virtually any opsin cDNAs can be expressed in cultured cells, reconstituted with 11-cis-retinal, and the absorption spectra of the resulting visual pigments can be measured. These advances in vision research also provide a rare opportunity for the study of adaptive evolution at the molecular level. The coelacanths (Latimeria chalumnae) live at a depth of 200 m near the coast of the Comoros Islands in the western Indian Ocean. The ocean floor at the depth of approximately 200 m receives only a narrow range of sunlight at approximately 480 nm. Out of the five groups of visual pigments, the coelacanths have retained only RH1 (African coelacanth; P485) and RH2 (African coelacanth; P478) pigments. Note that, compared with those of most orthologous pigments, the lmax of these two pigments are reduced by approximately 10–20 nm (Figure 2) and their absorption spectra have been devised to visualize the entire spectrum of color available to the coelacanths (Yokoyama et al., 1999). How did the coelacanths achieve these exquisitely coordinated blue shifts in the lmax of the two pigments? Comparative amino acid sequence analyses suggest that E122Q/A292S (amino acid changes E ? Q and A ? S at sites 122 and 292 respectively, and E122Q/M207L occurred along the branches leading to the coelacanth RH1 and RH2 pigments respectively. Indeed, amino acid changes Q122E/S292A in the RH1 pigment and Q122E/ L207M in the RH2 pigment increase the lmax by 26 and 21 nm respectively. Thus, the blue shift in the lmax in the RH1 pigment has been explained well by E122Q/A292S and that of the RH2 pigment by E122Q/M207L (Yokoyama et al., 1999). These amino acid sites are located in the transmembrane segments (Figure 3).

LWS and MWS Pigments Many MWS and LWS pigments have lmax of approximately 530 and 560 nm respectively (Figure 2). It can be shown experimentally that this 30 nm difference in the lmax is caused by amino acid

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differences at three sites: A164/F261/A269 in the MWS pigments and S164/Y261/T269 in the LWS pigments. Having LWS pigment-like amino acids A164/Y261/T269, however, the orthologous pigments in mouse, rat and rabbit have lmax of approximately 510 nm (Figure 2). These extremely blue-shifted lmax are shown to be achieved by H181Y/A292S. Thus, to explore the spectral tuning in the LWS/MWS pigments in all vertebrates, we should consider amino acid replacements at five sites: 164, 181, 261, 269 and 292. Multiple regression analyses based on these five sites of 26 currently known LWS/MWS pigments strongly suggest that the pigment in the vertebrate ancestor had an amino acid composition of S164/H181/Y261/T269/ A292 with a lmax of 559 nm and that mutations S164A, H181Y, Y261F, T269A, A292S and S164A/ H181Y in this ancestral pigment shift the lmax by 7, 28, 8, 15, 27 and 11 nm respectively (Yokoyama and Radlwimmer, 2001). These sites are located either in the transmembrane segments or very close to them (Figure 3). Importantly, extensive mutagenesis experiments also reveal that amino acid changes at the background sites do not cause any lmax shift, showing that the spectral tuning in the LWS/MWS pigments is explained fully by the ‘five-sites’ rule (Yokoyama and Radlwimmer, 2001). In higher primates, the LWS/MWS pigments evolved in two different ways. Hominoid and Old World monkeys use LWS and MWS opsins, which are encoded by two separate X-linked loci. Most New

164 83

C

211

122

86 52

261 265 269 295

49 46 90

292

93 118 114 181

N

207

Figure 3 Secondary structure of bovine RH1 opsin, showing naturally occurring amino acid mutations that cause more than 5 nm of lmax shift. The model is based on Palczewski et al. (2000). Open square, filled square and filled circles indicate the amino acid sites that are involved mainly in the spectral tuning of SWS1, LWS/ MWS and RH1/RH2 pigments respectively (see Yokoyama et al., 2000; Shi et al., 2001).


World monkeys, however, have one corresponding X-linked locus with three alleles (Figure 1). In these species, therefore, all males are so-called red–green color blind, whereas females are either ‘color blind’ or have complete red–green color vision depending on the genotype. In human, red–green color vision is controlled by LWS and MWS pigments. On the human X chromosome, one LWS and one or multiple MWS genes are located. Intragenic recombination between LWS and MWS genes produces both 50 MWS–LWS 30 and 50 LWS–MWS 30 hybrid genes (Nathans et al., 1986). The addition of the 50 MWS–LWS 30 gene to an otherwise normal gene array containing wild-type LWS and MWS genes causes the most common inherited color vision anomaly, deuteranomaly (Nathans et al., 1986). Many deuteranomalous men have wild-type MWS gene(s) that do not contribute to correct the anomalous color vision (Neitz et al., 1996). It should be noted that, as long as the exon 5 of the hybrid gene encodes Y261 and T269, the red vision is restored. Thus, if we define any LWS/MWS genes that encode Y261 and T269 as LWS genes, many human X chromosomes have more than one LWS gene, and sometimes even up to four (Neitz and Neitz, 1995). In human populations, it has been observed that approximately 60% of the LWS pigments have S164 and approximately 40% of the allelic LWS pigments have A164 (Winderickx et al., 1992). Among MWS genes, variation at site 164 is less common, as at least 90% encode A164 (Nathans, 1999). The amino acid dimorphism can cause 7 nm difference in the lightsensitivities of the two types of LWS pigments.

SWS1 Pigments Many fishes, amphibians, reptiles, birds and some mammals use ultraviolet (UV) vision for such basic activities as foraging and mate choice. These species detect light maximally at 360–370 nm by using UV pigments. These UV pigments and violet (or blue) pigments with lmax of 390–430 nm belong to the same SWS1 group (Figure 2). The spectral tuning in the UV pigments has been studied first by considering avian pigments, and then mammalian pigments. The zebra finch, blackbird, canary and budgerigar SWS1 pigments have lmax of 358–366 nm, whereas the orthologous violet pigments of pigeon and chicken have lmax of more than 390 nm (Figure 2). It has been shown that the avian UV pigments evolved from the violet pigment by one amino acid replacement, S90C (Wilkie et al., 2000; Yokoyama et al., 2000). On the other hand, the mouse SWS1 pigment has a lmax of 359 nm, whereas the closely related human blue pigment has a lmax of 414 nm. The mouse UV pigment can be made

blue-sensitive (lmax ¼ 411 nm) by introducing seven amino acid changes F46T/F49L/T52F/F86L/T93P/ A114G/S118T, whereas the human blue pigment can be made into UV pigment with a lmax of 360 nm by introducing the seven reverse mutations (Shi et al., 2001). These analyses show that the violet pigments evolved from the UV pigment by accumulating at least two of the eight amino acid replacements. However, F86Y had the major impact in shifting the lmax by more than 50 nm (Fasick et al., 2002). These results suggest that the difference between the UV and violet pigments in vertebrates is based on a total of eight amino acid sites 46, 49, 52, 86, 90, 93, 114 and 118, which are all located in the transmembrane segments (Figure 3). Comparative amino acid sequence analyses suggest that the common ancestral SWS1 pigment in vertebrates had amino acids F46/ F49/T52/F86/S90/T93/A114/S118. This amino acid composition is identical to those of the contemporary salamander, chameleon, mouse and rat UV pigments with lmax of approximately 360 nm (Figure 2), but T93Q occurred in the ancestral fish UV pigment. Using the goldfish UV pigment, it has been shown that Q93T does not shift the lmax from that of the wild-type pigment. Thus, the ancestral pigment in vertebrates must have had a lmax of approximately 360 nm, and the fish, salamander, chameleon, mouse and rat pigments have maintained their UV sensitivities through purifying selection (Shi et al., 2001). In the avian lineage, the ancestral pigment lost UV sensitivity, but some descendants regained it by S90C. Because of the nonadditive effects of amino acid changes on the lmax shift, the evolutionary processes of the functional differentiation of various violet pigments remain to be elucidated.

Perspectives The comparative sequence analyses followed by the mutagenesis experiments demonstrate that the evolutionary approach is a powerful method in enhancing our understanding of the functional differentiations of a wide variety of visual pigment genes. As more amino acid sequences and absorption spectra of visual pigments accumulate, the prediction of potentially important amino acid changes in the spectral tuning in visual pigments will become more accurate. Sampling of visual pigments from various photic environments or those associated with different behavioral characteristics would be of particular interest, because we may also uncover previously unknown amino acid sites that are involved in the spectral tuning in visual pigments. Visual pigments associated with specific photic environments or unique behaviors also provide

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an excellent opportunity to analyze adaptive evolution at the molecular level.

See also Color Vision Defects Eye: Proteomics

References Fasick JI, Applebury ML and Oprian DD (2002) Spectral tuning in the mammalian short-wavelength sensitive cone pigments. Biochemistry 41: 6860–6865. Nathans J (1999) The evolution and physiology of human color vision: insights from molecular genetic studies of visual pigments. Neuron 24: 299–312. Nathans J, Thomas D and Hogness DS (1986) Molecular genetics of human color vision: the genes encoding blue, green, and red pigments. Science 232: 193–201. Neitz M and Neitz J (1995) Numbers and ratios of visual pigment genes for normal red–green color vision. Science 267: 1013–1016. Neitz J, Neitz M and Kainz PM (1996) Visual pigment gene structure and the severity of color vision defects. Science 274: 801–803. Palczewski K, Kumasaka T, Hori T, et al. (2000) Crystal structure of rhodopsin: a G protein-coupled receptor. Science 289: 739–745. Saitou N and Nei M (1987) The neighbor-joining method: a new method for estimating phylogenetic trees. Molecular Biology and Evolution 4: 406–425. Shi Y, Radlwimmer FB and Yokoyama S (2001) Molecular genetics and the evolution of ultraviolet vision in vertebrates. Proceedings of the National Academy of Sciences of the United States of America 98: 11 731–11 736. Wilkie SE, Robinson PR, Cronin TW, et al. (2000) Spectral tuning of avian violet- and ultraviolet-sensitive visual pigments. Biochemistry 39: 7895–7901.

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Winderickx J, Lindsey DT, Sanocki E, et al. (1992) Polymorphism in red photopigment underlies variation in colour matching. Nature 356: 431–433. Yokoyama S (2000) Molecular evolution of vertebrate visual pigments. Progress in Retinal and Eye Research 19: 385–419. Yokoyama S and Radlwimmer FB (2001) The molecular genetics and evolution of red and green color vision in vertebrates. Genetics 158: 1697–1710. Yokoyama S, Radlwimmer FB and Blow NS (2000) Ultraviolet pigments in birds evolved from violet pigments by a single amino acid change. Proceedings of National Academy of Sciences of the United States of America 97: 7366–7371. Yokoyama S, Zhang H, Radlwimmer FB and Blow NS (1999) Adaptive evolution of color vision of the Comoran coelacanth (Latimeria chalumnae). Proceedings of National Academy of Sciences of the United States of America 96: 6279–6284.

Further Reading Ebrey T and Koutalos Y (2001) Vertebrate photoreceptors. Progress in Retinal and Eye Research 20: 49–94. Kochendoerfer GG, Lin SW, Sakmar TP and Mathies RA (1999) How color visual pigments are tuned. Trends in Biochemical Sciences 24: 300–305. Nathans J (1990) Determinants of visual pigment absorbance: role of changed amino acids in the putative transmembrane segments. Biochemistry 29: 937–942. Sharpe LT, Stockman A, Jagle H, et al. (1998) Red, green and red–green hybrid pigments in the human retina: correlations between deduced protein sequences and psychophysically measured spectral sensitivities. Journal of Neuroscience 18: 10 053–10 069. Yokoyama S (2002) Molecular evolution of color vision in vertebartes. Gene 300: 69–78.