Diversity of color vision: not all Australian marsupials are trichromatic

Color vision in marsupials has recently emerged as a particularly interesting case among mammals. It appears that there are both dichromats and trichromats among closely related species. In contrast to primates, marsupials seem to have evolved a different type of trichromacy that is not linked to the X-chromosome. Based on microspectrophotometry and retinal whole-mount immunohistochemistry, four trichromatic marsupial species have been described: quokka, quenda, honey possum, and fat-tailed dunnart. It has, however, been impossible to identify the photopigment of the third cone type, and genetically, all evidence so far suggests that all marsupials are dichromatic. The tammar wallaby is the only Australian marsupial to date for which there is no evidence of a third cone type. To clarify whether the wallaby is indeed a dichromat or trichromatic like other Australian marsupials, we analyzed the number of cone types in the "dichromatic" wallaby and the "trichromatic" dunnart. Employing identical immunohistochemical protocols, we confirmed that the wallaby has only two cone types, whereas 20-25% of cones remained unlabeled by S- and LM-opsin antibodies in the dunnart retina. In addition, we found no evidence to support the hypothesis that the rod photopigment (rod opsin) is expressed in cones which would have explained the absence of a third cone opsin gene. Our study is the first comprehensive and quantitative account of color vision in Australian marsupials where we now know that an unexpected diversity of different color vision systems appears to have evolved.     

Evolution and spectral tuning of visual pigments in birds and mammals

Variation in the types and spectral characteristics of visual pigments is a common mechanism for the adaptation of the vertebrate visual system to prevailing light conditions. The extent of this diversity in mammals and birds is discussed in detail in this review, alongside an in-depth consideration of the molecular changes involved. In mammals, a nocturnal stage in early evolution is thought to underlie the reduction in the number of classes of cone visual pigment genes from four to only two, with the secondary loss of one of these genes in many monochromatic nocturnal and marine species. The trichromacy seen in many primates arises from either a polymorphism or duplication of one of these genes. In contrast, birds have retained the four ancestral cone visual pigment genes, with a generally conserved expression in either single or double cone classes. The loss of sensitivity to ultraviolet (UV) irradiation is a feature of both mammalian and avian visual evolution, with UV sensitivity retained among mammals by only a subset of rodents and marsupials. Where it is found in birds, it is not ancestral but newly acquired.     

Functional diversity in the color vision of cichlid fishes

Background  

Color vision plays a critical role in visual behavior. An animal's capacity for color vision rests on the presence of differentially sensitive cone photoreceptors. Spectral sensitivity is a measure of the visual responsiveness of these cones at different light wavelengths. Four classes of cone pigments have been identified in vertebrates, but in teleost fishes, opsin genes have undergone gene duplication events and thus can produce a larger number of spectrally distinct cone pigments. In this study, we examine the question of large-scale variation in color vision with respect to individual, sex and species that may result from differential expression of cone pigments. Cichlid fishes are an excellent model system for examining variation in spectral sensitivity because they have seven distinct cone opsin genes that are differentially expressed.     

Cone topography and spectral sensitivity in two potentially trichromatic marsupials, the quokka (Setonix brachyurus) and quenda (Isoodon obesulus)

The potential for trichromacy in mammals, thought to be unique to primates, was recently discovered in two Australian marsupials. Whether the presence of three cone types, sensitive to short- (SWS), medium- (MWS) and long- (LWS) wavelengths, occurs across all marsupials remains unknown. Here, we have investigated the presence, distribution and spectral sensitivity of cone types in two further species, the quokka (Setonix brachyurus) and quenda (Isoodon obesulus). Immunohistochemistry revealed that SWS cones in the quokka are concentrated in dorso-temporal retina, while in the quenda, two peaks were identified in naso-ventral and dorso-temporal retina. In both species, MWS/LWS cone spatial distributions matched those of retinal ganglion cells. Microspectrophotometry (MSP) confirmed that MWS and LWS cones are spectrally distinct, with mean wavelengths of maximum absorbance at 502 and 538 nm in the quokka, and at 509 and 551 nm, in the quenda. Although small SWS cone outer segments precluded MSP measurements, molecular analysis identified substitutions at key sites, accounting for a spectral shift from ultraviolet in the quenda to violet in the quokka. The presence of three cone types, along with previous findings in the fat-tailed dunnart and honey possum, suggests that three spectrally distinct cone types are a feature spanning the marsupials.     

Bat Eyes Have Ultraviolet-Sensitive Cone Photoreceptors

Mammalian retinae have rod photoreceptors for night vision and cone photoreceptors for daylight and colour vision. For colour discrimination, most mammals possess two cone populations with two visual pigments (opsins) that have absorption maxima at short wavelengths (blue or ultraviolet light) and long wavelengths (green or red light). Microchiropteran bats, which use echolocation to navigate and forage in complete darkness, have long been considered to have pure rod retinae. Here we use opsin immunohistochemistry to show that two phyllostomid microbats, Glossophaga soricina and Carollia perspicillata, possess a significant population of cones and express two cone opsins, a shortwave-sensitive (S) opsin and a longwave-sensitive (L) opsin. A substantial population of cones expresses S opsin exclusively, whereas the other cones mostly coexpress L and S opsin. S opsin gene analysis suggests ultraviolet (UV, wavelengths <400 nm) sensitivity, and corneal electroretinogram recordings reveal an elevated sensitivity to UV light which is mediated by an S cone visual pigment. Therefore bats have retained the ancestral UV tuning of the S cone pigment. We conclude that bats have the prerequisite for daylight vision, dichromatic colour vision, and UV vision. For bats, the UV-sensitive cones may be advantageous for visual orientation at twilight, predator avoidance, and detection of UV-reflecting flowers for those that feed on nectar.     

Why Aye‐Ayes See Blue

The capacity for cone‐mediated color vision varies among nocturnal primates. Some species are colorblind, having lost the functionality of their short‐wavelength‐sensitive‐1 (SWS1) opsin pigment gene. In other species, such as the aye‐aye (Daubentonia madagascariensis), the SWS1 gene remains intact. Recent studies focused on aye‐ayes indicate that this gene has been maintained by natural selection and that the pigment has a peak sensitivity (λ_max) of 406 nm, which is ～20 nm closer to the ultraviolet region of the spectrum than in most primates. The functional significance behind the retention and unusual λ_max of this opsin pigment is unknown, and it is perplexing given that all mammals are presumed to be colorblind in the dark. Here we comment on this puzzle and discuss recent findings on the color vision intensity thresholds of terrestrial vertebrates with comparable optics to aye‐ayes. We draw attention to the twilight activities of aye‐ayes and report that twilight is enriched in short‐wavelength (bluish) light. We also show that the intensity of twilight and full moonlight is probably sufficient to support cone‐mediated color vision. We speculate that the intact SWS1 opsin pigment gene of aye‐ayes is a crepuscular adaptation and we report on the blueness of potential visual targets, such as scent marks and the brilliant blue arils of Ravenala madagascariensis.     

Isolation and characterization of melanopsin (Opn4) from the Australian marsupial Sminthopsis crassicaudata (fat-tailed dunnart)

Melanopsin confers photosensitivity to a subset of retinal ganglion cells and is responsible for many non-image-forming tasks, like the detection of light for circadian entrainment. Recently, two melanopsin genes, Opn4m and Opn4x, were described in non-mammalian vertebrates. However, only one form, Opn4m, has been described in the mammals, although studies to date have been limited to the placentals and have not included the marsupials. We report here the isolation and characterization of an Opn4 gene from an Australian marsupial, the fat-tailed dunnart (Sminthopsis crassicaudata), and present evidence which suggests that the Opn4x gene was lost before the placental/marsupial split. In situ hybridization shows that the expression of Opn4 in the dunnart eye is restricted to a subset of ganglion cells, a pattern previously reported for rodents and primates. These Opn4-positive cells are randomly distributed across the dunnart retina. We also undertook a comparative analysis with the South American marsupial, the grey short-tailed opossum (Monodelphis domestica), and two placental mammals, mouse and human. This approach reveals that the two marsupials show a higher sequence identity than that seen between rodents and primates, despite separating at approximately the same point in time, some 65-85 Myr ago.     

Diurnality and cone photopigment polymorphism in strepsirrhines: examination of linkage in Lemur catta

Trichromatic color vision is routine among catarrhine primates, but occurs only as a variant form of color vision in some individuals in most platyrrhine genera. This arises from a fundamental difference in the organization of X-chromosome cone opsin genes in these two lineages: catarrhines have two opsin genes specifying middle- and long-wavelength-sensitive cone pigments, while platyrrhines have only a single gene. Some female platyrrhine monkeys achieve trichromacy because of a species polymorphism that allows the possibility of different opsin gene alleles on the two X-chromosomes. Recently, a similar opsin gene polymorphism was detected in some diurnal strepsirrhines, while at the same time appearing to be absent in any nocturnal genera. The aim of this study was to assess whether cone pigment polymorphism is inevitably linked to diurnality in strepsirrhines. Cone photopigments were measured in a species usually classified as diurnal, the ring-tailed lemur (Lemur catta), using electroretinogram flicker photometry, a noninvasive electrophysiological procedure. Each of 12 animals studied was found to have the same middle-wavelength cone pigment, with peak sensitivity at about 547 nm. In conjunction with earlier results, this implies that cone pigment polymorphism is unlikely to exist in this species and that, accordingly, such variation is not a consistently predictable feature of vision in diurnal strepsirrhines.     

Platypus TCRμ provides insight into the origins and evolution of a uniquely mammalian TCR locus

TCRμ is an unconventional TCR that was first discovered in marsupials and appears to be absent from placental mammals and nonmammals. In this study, we show that TCRμ is also present in the duckbill platypus, an egg-laying monotreme, consistent with TCRμ being ancient and present in the last common ancestor of all extant mammals. As in marsupials, platypus TCRμ is expressed in a form containing double V domains. These V domains more closely resemble Ab V than that of conventional TCR. Platypus TCRμ differs from its marsupial homolog by requiring two rounds of somatic DNA recombination to assemble both V exons and has a genomic organization resembling the likely ancestral form of the receptor genes. These results demonstrate that the ancestors of placental mammals would have had TCRμ but it has been lost from this lineage.     

Vision in elasmobranchs and their relatives: 21st century advances

This review identifies a number of exciting new developments in the understanding of vision in cartilaginous fishes that have been made since the turn of the century. These include the results of studies on various aspects of the visual system including eye size, visual fields, eye design and the optical system, retinal topography and spatial resolving power, visual pigments, spectral sensitivity and the potential for colour vision. A number of these studies have covered a broad range of species, thereby providing valuable information on how the visual systems of these fishes are adapted to different environmental conditions. For example, oceanic and deep-sea sharks have the largest eyes amongst elasmobranchs and presumably rely more heavily on vision than coastal and benthic species, while interspecific variation in the ratio of rod and cone photoreceptors, the topographic distribution of the photoreceptors and retinal ganglion cells in the retina and the spatial resolving power of the eye all appear to be closely related to differences in habitat and lifestyle. Multiple, spectrally distinct cone photoreceptor visual pigments have been found in some batoid species, raising the possibility that at least some elasmobranchs are capable of seeing colour, and there is some evidence that multiple cone visual pigments may also be present in holocephalans. In contrast, sharks appear to have only one cone visual pigment. There is evidence that ontogenetic changes in the visual system, such as changes in the spectral transmission properties of the lens, lens shape, focal ratio, visual pigments and spatial resolving power, allow elasmobranchs to adapt to environmental changes imposed by habitat shifts and niche expansion. There are, however, many aspects of vision in these fishes that are not well understood, particularly in the holocephalans. Therefore, this review also serves to highlight and stimulate new research in areas that still require significant attention.     

Generation of knock-in mice carrying third cones with spectral sensitivity different from S and L cones

Red-green color vision in primates is unique in the sense that it is mediated by two photoreceptor cells that are indistinguishable in all aspects except for their visual pigments. In order to generate an animal model for investigation of the interaction between red-green inputs at the molecular level, we applied knock-in technology and X-chromosome inactivation machinery to make a mouse model with cone cells possessing visual pigments with different spectral sensitivities. We introduced a S308A point mutation into the Green opsin gene allele on the X-chromosome. This manipulation generated a 24 nm red-shift of absorption maximum in the cone pigment with negligible functional differences in other molecular properties. Amplitudes of responses in ERG and ganglion cell recordings of homozygotes were similar to those of wild-types, although the spectral sensitivities differed. Heterozygotes showed variable spectral sensitivities of ganglion cell responses due to the different integration of the native and the S308A cone inputs on the dendritic fields. In situ hybridization experiments showed that cone cells with respective pigments formed patch-like clusters of specific L cone-types, approximately 30 mum in diameter, which were randomly distributed in the dorsal region of the retinas. Since the patch-like clustering was arranged by X-inactivation, such clustering could be present in the peripheral retinas of New World monkeys with polymorphic L pigments, indicating that our mice would be a suitable model to study evolution of the mammalian color vision system.     

Molecular Evolution of Arthropod Color Vision Deduced from Multiple Opsin Genes of Jumping Spiders

Among terrestrial animals, only vertebrates and arthropods possess wavelength-discrimination ability, so-called “color vision”. For color vision to exist, multiple opsins which encode visual pigments sensitive to different wavelengths of light are required. While the molecular evolution of opsins in vertebrates has been well investigated, that in arthropods remains to be elucidated. This is mainly due to poor information about the opsin genes of non-insect arthropods. To obtain an overview of the evolution of color vision in Arthropoda, we isolated three kinds of opsins, Rh1, Rh2, and Rh3, from two jumping spider species, Hasarius adansoni and Plexippus paykulli. These spiders belong to Chelicerata, one of the most distant groups from Hexapoda (insects), and have color vision as do insects. Phylogenetic analyses of jumping spider opsins revealed a birth and death process of color vision evolution in the arthropod lineage. Phylogenetic positions of jumping spider opsins revealed that at least three opsins had already existed before the Chelicerata-Pancrustacea split. In addition, sequence comparison between jumping spider Rh3 and the shorter wavelength-sensitive opsins of insects predicted that an opsin of the ancestral arthropod had the lysine residue responsible for UV sensitivity. These results strongly suggest that the ancestral arthropod had at least trichromatic vision with a UV pigment and two visible pigments. Thereafter, in each pancrustacean and chelicerate lineage, the opsin repertoire was reconstructed by gene losses, gene duplications, and function-altering amino acid substitutions, leading to evolution of color vision. Mitsumasa Koyanagi and Takashi Nagata contributed equally to this work. Sequence data from this article have been deposited with the DDBJ under accession nos. AB251846–AB251851.     

Mix and match color vision: tuning spectral sensitivity by differential opsin gene expression in Lake Malawi cichlids

Cichlid fish of the East African Rift Lakes are renowned for their diversity and offer a unique opportunity to study adaptive changes in the visual system in rapidly evolving species flocks. Since color plays a significant role in mate choice, differences in visual sensitivities could greatly influence and even drive speciation of cichlids. Lake Malawi cichlids inhabiting rock and sand habitats have significantly different cone spectral sensitivities. By combining microspectrophotometry (MSP) of isolated cones, sequencing of opsin genes, and spectral analysis of recombinant pigments, we have established the cone complements of four species of Malawi cichlids. MSP demonstrated that each of these species predominately expresses three cone pigments, although these differ between species to give three spectrally different cone complements. In addition, rare populations of spectrally distinct cones were found. In total, seven spectral classes were identified. This was confirmed by opsin gene sequencing, expression, and in vitro reconstitution. The genes represent the four major classes of cone opsin genes that diverged early in vertebrate evolution. All four species possess a long-wave-sensitive (LWS), three spectrally distinct green-sensitive (RH2), a blue-sensitive (SWS2A), a violet-sensitive (SWS2B), and an ultraviolet-sensitive (SWS1) opsin. However, African cichlids determine their spectral sensitivity by differential expression of primarily only three of the seven available cone opsin genes. Phylogenetic analysis suggests that all percomorph fish have similar potential.     

Fruits, foliage and the evolution of primate colour vision

Primates are apparently unique amongst the mammals in possessing trichromatic colour vision. However, not all primates are trichromatic. Amongst the haplorhine (higher) primates, the catarrhines possess uniformly trichromatic colour vision, whereas most of the platyrrhine species exhibit polymorphic colour vision, with a variety of dichromatic and trichromatic phenotypes within the population. It has been suggested that trichromacy in primates and the reflectance functions of certain tropical fruits are aspects of a coevolved seed-dispersal system: primate colour vision has been shaped by the need to find coloured fruits amongst foliage, and the fruits themselves have evolved to be salient to primates and so secure dissemination of their seeds. We review the evidence for and against this hypothesis and we report an empirical test: we show that the spectral positioning of the cone pigments found in trichromatic South American primates is well matched to the task of detecting fruits against a background of leaves. We further report that particular trichromatic platyrrhine phenotypes may be better suited than others to foraging for particular fruits under particular conditions of illumination; and we discuss possible explanations for the maintenance of polymorphic colour vision amongst the platyrrhines.     

Comparative locomotor performance of marsupial and placental mammals

Marsupials are often considered inferior to placental mammals in a number of physiological characters. Because locomotor performance is presumed to be an important component of fitness, we compared marsupials and placentals with regard to both maximal running speeds and maximal aerobic speeds (=speed at which the maximal rate of oxygen consumption, VOlmax, is attained). Maximal aerobic speed is related to an animal's maximal sustainable speed, and hence is a useful comparative index of stamina.
Maximal running speeds of 11 species of Australian marsupials, eight species of Australian murid rodents, two species of American didelphid marsupials, and two species of American rodents were measured in the laboratory and compared with data compiled from the literature. Our values are greater than, or equivalent to, those reported previously. Marsupials and placentals do not differ in maximal running speeds (nor do Australian rodents differ from non-Australian rodents). Within these groups, however, species and families may differ considerably. Some of the interspecific variation in maximal running speeds is related to differences in habitat: species inhabiting open habitats (e.g. deserts) tend to be faster than are species from habitats with more cover, or arboreal species.
Maximal aerobic speeds (compiled from the literature) were higher in large species than in small species. However, marsupials and placentals show no general difference with regard to maximal aerobic speeds.
Maximal running speeds and maximal aerobic speeds for 18 species of mammals were not correlated, after correcting for correlations with body size. Thus, the fastest sprinters do not necessarily have high maximal aerobic speeds.     

Genome sequence comparison reveals independent inactivation of the caspase-15 gene in different evolutionary lineages of mammals

We have recently demonstrated that placental mammalian species such as pig and dog express a novel proapoptotic protease, caspase-15, whereas mouse and humans lack this enzyme. Here we investigated the evolutionary fate of the caspase-15 gene in different mammalian lineages by analyzing whole-genome shotgun sequences of 30 mammalian species for the presence of caspase-15 orthologs. Caspase-15 gene sequences were found in representatives of all major mammalian clades except for the superorders Afrotheria (tenrec, rock hyrax, and elephant) and Euarchontoglires (rodents, rabbit, tree shrew, and primates), which either lacked any caspase-15-like sequences or contained mutated remnants of the caspase-15 gene. Polymerase chain reaction screenings confirmed the results of the database searches and showed that the caspase-15 gene is expressed not only in various placental mammals but also in the marsupial, Monodelphis domestica. The observed species distribution implies that caspase-15 has originated in an early ancestor of modern mammals and has been conserved, over more than 180 Myr, in marsupials and many placental mammals, whereas it was independently lost in 2 phylogenetically distant clades of placental mammals, that is, Afrotheria and Euarchontoglires. Our data suggest that the inactivation of the caspase-15 gene was not counteracted by, and may even have been driven by, evolutionary constraints in these clades, and therefore, caution against the uncritical use of gene absence for the inference of phylogenetic relationships.     

Dual duplication of neurohypophysial hormones in an Australian marsupial: mesotocin, oxytocin, lysine vasopressin and arginine vasopressin in a single gland of the northern bandicoot (Isoodon macrourus)

Neurohypophysial hormones of an Australian marsupial, the Northern bandicoot (Isoodon macrourus), have been identified by their retention times in high-pressure reverse-phase liquid chromatography using two solvent systems and by their molar pressor or uterotonic activities. Two pressor peptides, arginine vasopressin and lysipressin, and two uterotonic peptides, mesotocin and oxytocin, have been characterized. Because mesotocin and arginine vasopressin have been identified in three other Australian marsupial families, it is assumed that a duplication of each ancestral gene occurred in Peramelidae and subsequent mutations in one copy led to the additional oxytocin and lysipressin. A similar dual duplication of neurohypophysial hormones has previously been discovered in the North-American opossum (Didelphis virginiana) so that the duplication propensity seems peculiar to marsupials in contrast to placental mammals.     

The cone visual pigments of an Australian marsupial,the tammar wallaby (Macropus eugenii): sequence,spectral tuning,and evolution

Studies on marsupial color vision have been limited to very few species. There is evidence from behavioral, electroretinographic (ERG), and microspectrophotometric (MSP) measurements for the existence of both dichromatic and trichromatic color vision. No studies have yet investigated the molecular mechanisms of spectral tuning in the visual pigments of marsupials. Our study is the first to determine the mRNA sequence, infer the amino acid sequence, and determine, by in vitro expression, the spectra of the cone opsins of a marsupial, the tammar wallaby (Macropus eugenii). This yielded some information on mechanisms and evolution of spectral tuning of these pigments. The tammar wallaby retina contains only short-wavelength sensitive (SWS) and middle-wavelength sensitive (MWS) pigment mRNAs. This predicts dichromatic color vision, which is consistent with conclusions from previous behavioral studies ( Hemmi 1999). We found that the wallaby has a SWS1 class pigment of 346 amino acids. Sequence comparison with eutherian SWS pigments predicts that this SWS1 pigment absorbs maximally (lambdamax) at 424 nm and, therefore, is a blue rather than a UV pigment. This (lambdamax) is close to that of the in vitro-expressed wallaby SWS pigment (lambdamax of 420 +/- 2 nm) and to that determined behaviorally (420 nm). The difference from the mouse UV pigment (lambdamax of 359 nm) is largely accounted for by the F86Y substitution, in agreement with in vitro results comparing a variety of other SWS pigments. This suggests that spectral tuning employing F86Y substitution most likely arose independently in the marsupials and ungulates as a result of convergent evolution. An apparently different mechanism of spectral tuning of the SWS1 pigments, involving five amino acid positions, evolved in primates. The wallaby MWS pigment has 363 amino acids. Species comparisons at positions critical to spectral tuning predict a lambdamax near 530 nm, which is close to that of the in vitro-expressed pigment (529 +/- 1 nm), but quite different from the value of 539 nm determined by microspectrophotometry. Introns interrupt the coding sequences of the wallaby, mouse, and human MWS pigment sequences at the same corresponding nucleotide positions. However, the length of introns varies widely among these species.     

Molecular characterization of visual pigments in Branchiopoda and the evolution of opsins in Arthropoda

Studies on color vision in invertebrates have focused primarily on insect visual pigments, with little attention given to crustacean visual pigments. None of the blue-green-, blue-, or ultraviolet (UV)-sensitive-opsins have been identified in crustaceans. In addition, the discussion of visual pigments has been limited to long-wavelength-sensitive opsins in Pancrustacea. Here, we focused on Branchiopoda (Crustacea), which is a sister group of Hexapoda including insects. In the tadpole shrimp Triops granarius, the visual pigment chromophore was retinal. Multiple opsins were isolated from each of three branchiopod species, T. granarius, Triops longicaudatus, and the fairy shrimp Branchinella kugenumaensis (five, five, and four opsins from these species, respectively). Phylogenetic analyses and the presence of a lysine residue corresponding to position 90 in bovine rhodopsin suggested that three of the branchiopod opsins comprise UV-sensitive pigments. In addition, the phylogenetic relationships between insect and branchiopod UV-sensitive opsins revealed that the divergence of blue- and UV-sensitive pigments predates the Branchiopoda and Insecta divergence. The other branchiopod opsins show distant relationships to other known insect opsins and form novel clusters. The present results strongly suggest that the ancestral arthropod of the Chelicerata-Pancrustacea lineages possessed at least four types of opsins. The ancestors of Pancrustacea and the Insecta-Branchiopoda lineages possessed at least five and six types of opsins, respectively. Our results suggest that in the evolutionary process associated with each lineage, several opsins appeared and diversified with repeated gene duplication, of which some have been lost in some taxa.