The molecular genetics of red and green color vision in mammals.

To elucidate the molecular mechanisms of red-green color vision in mammals, we have cloned and sequenced the red and green opsin cDNAs of cat (Felis catus), horse (Equus caballus), gray squirrel (Sciurus carolinensis), white-tailed deer (Odocoileus virginianus), and guinea pig (Cavia porcellus). These opsins were expressed in COS1 cells and reconstituted with 11-cis-retinal. The purified visual pigments of the cat, horse, squirrel, deer, and guinea pig have lambdamax values at 553, 545, 532, 531, and 516 nm, respectively, which are precise to within +/-1 nm. We also regenerated the "true" red pigment of goldfish (Carassius auratus), which has a lambdamax value at 559 +/- 4 nm. Multiple linear regression analyses show that S180A, H197Y, Y277F, T285A, and A308S shift the lambdamax values of the red and green pigments in mammals toward blue by 7, 28, 7, 15, and 16 nm, respectively, and the reverse amino acid changes toward red by the same extents. The additive effects of these amino acid changes fully explain the red-green color vision in a wide range of mammalian species, goldfish, American chameleon (Anolis carolinensis), and pigeon (Columba livia).     

The molecular genetics and evolution of red and green color vision in vertebrates.

To better understand the evolution of red-green color vision in vertebrates, we inferred the amino acid sequences of the ancestral pigments of 11 selected visual pigments: the LWS pigments of cave fish (Astyanax fasciatus), frog (Xenopus laevis), chicken (Gallus gallus), chameleon (Anolis carolinensis), goat (Capra hircus), and human (Homo sapiens);and the MWS pigments of cave fish, gecko (Gekko gekko), mouse (Mus musculus), squirrel (Sciurus carolinensis), and human. We constructed these ancestral pigments by introducing the necessary mutations into contemporary pigments and evaluated their absorption spectra using an in vitro assay. The results show that the common ancestor of vertebrates and most other ancestors had LWS pigments. Multiple regression analyses of ancestral and contemporary MWS and LWS pigments show that single mutations S180A, H197Y, Y277F, T285A, A308S, and double mutations S180A/H197Y shift the lambda(max) of the pigments by -7, -28, -8, -15, -27, and 11 nm, respectively. It is most likely that this "five-sites" rule is the molecular basis of spectral tuning in the MWS and LWS pigments during vertebrate evolution.     

The evolution of color vision in the primates

There will be no difficulty in seeing how and by what mixtures the colors are made … He, however, who should attempt to verify all this by experiment would forget the difference of the human and the divine nature. For God only has the knowledge and also the power which are able to combine many things into one and again resolve the one into many. But no man either is or ever will be able to accomplish either the one or the other operation.The law of proportion according to which the several colors are formed, even if a man knew he would be foolish in telling, for he could not give any necessary reason, nor indeed any tolerable or probable explanation of them (Jowett, 1871).     

Seeing red: color vision in the largemouth bass

How animals visually perceive the environment is key to understanding important ecological behaviors, such as predation, foraging, and mating. This study focuses on the visual system properties and visual perception of color in the largemouth bass Micropterus salmoides. This study (1) documents the number and spectral sensitivity of photoreceptors,(2) uses these parameters to model visual perception, and (3) tests the model of color perception using a behavioral assay. Bass possess single cone cells maximally sensitive at 535 nm, twin cone cells maximally sensitive at 614 nm, and rod cells maximally sensitive at 528 nm. A simple model of visual perception predicted that bass should not be able to discern between chartreuse yellow and white nor between green and blue. In contrast, bass should be able to discern red from all achromatic (i.e., gray scale) stimuli. These predictions were partially upheld in behavioral trials. In behavioral trials, bass were first trained to recognize a target color to receive a food reward, and then tested on their ability to differentiate between their target color and a color similar in brightness. Bass trained to red and green could easily discern their training color from all other colors for target colors that were similar in brightness (white and black, respectively). This study shows that bass possess dichromatic vision and do use chromatic (i.e., color) cues in making visual-based decisions.     

Historical contingency in the evolution of primate color vision

Primates are unique among eutherian mammals for possessing three types of retinal cone. Curiously, catarrhines, platyrrhines, and strepsirhines share this anatomy to different extents, and no hypothesis has hitherto accounted for this variability. Here we propose that the historical biogeography of figs and arborescent palms accounts for the global variation in primate color vision. Specifically, we suggest that primates invaded Paleogene forests characterized by figs and palms, the fruits of which played a keystone function. Primates not only relied on such resources, but also provided high-quality seed dispersal. In turn, figs and palms lost or simply did not evolve conspicuous coloration, as this conferred little advantage for attracting mammals. We suggest that the abundance and coloration of figs and palms offered a selective advantage to foraging groups with mixed capabilities for chromatic distinction. Climatic cooling at the end of the Eocene and into the Neogene resulted in widespread regional extinction or decimation of palms and (probably) figs. In regions where figs and palms became scarce, we suggest primates evolved routine trichromatic vision in order to exploit proteinaceous young leaves as a replacement resource. A survey of the hue and biogeography of extant figs and palms provides some empirical support. Where these resources are infrequent, primates are routinely trichromatic and consume young leaves during seasonal periods of fruit dearth. These results imply a link between the differential evolution of primate color vision and climatic changes during the Eocene-Oligocene transition.     

Molecular evolution of bat color vision genes

The two suborders of bats, Megachiroptera (megabats) and Microchiroptera(microbats), use different sensory modalities for perceivingtheir environment. Megabats are crepuscular and rely on a well-developedeyes and visual pathway, whereas microbats occupy a nocturnalniche and use acoustic orientation or echolocation more thanvision as the major means of perceiving their environment. Inview of the differences associated with their sensory systems,we decided to investigate the function and evolution of colorvision (opsin genes) in these two suborders of bats. The middle/longwavelength (M/L) and short wavelength (S) opsin genes were sequencedfrom two frugivorous species of megabats, Haplonycteris fischeriand Pteropus dasymallus formosus, and one insectivorous speciesof microbat, Myotis velifer. Contrary to the situation in primates,where many nocturnal species have lost the functional S opsingene, both crepuscular and strictly nocturnal species of batsthat we examined have functional M/L and S opsin genes. Surprisingly,the S opsin in these bats may be sensitive to UV light, whichis relatively more abundant at dawn and at dusk. The M/L opsinin these bats appears to be the L type, which is sensitive tored and may be helpful for identifying fruits among leaves orfor other purposes. Most interestingly, H. fischeri has a recentduplication of the M/L opsin gene, representing to date theonly known case of opsin gene duplication in non-primate mammals.Some of these observations are unexpected and may provide insightsinto the effect of nocturnal life on the evolution of opsingenes in mammals and the evolution of the life history traitsof bats in general.     

Use of the "SMURF" in taste analysis

An improved moving chart recording of intensity/time of tasteresponse has been achieved using a potentiometer ‘dialbox’ linked by a cable to a Telsec recorder. The deviceallows rates of taste response to be determined and is describedas a Sensory Measuring Unit for Recording Flux (SMURF) on theassumption that the flux of stimuli at the taste receptor isresponsible for the time course of response. Fourteen trained and sixteen untrained panellists evaluatedone standard and four unknown sucrose solutions using the SMURFand determined their intensity and persistence time of responsefor each of these same solutions by conventional interval scalingand use of a stop-clock. The SMURF gave results which were higher(but not significantly so) than the conventional method. Trainedpanellists tended to prefer the SMURF and found it quicker andeasier to use than the conventional method. Untrained panelliststended to prefer the conventional method but these results weregenerally not significant. The SMURF is therefore an extremelyuseful device in reducing time and effort whilst still maintainingaccuracy in the measurement of intensity and time of taste response. The SMURF was also used to obtain intensity/time data for threeother sugars so that a comparison between the sugars could bemade.     

The "Lost Year" Question in Young Sea Turtles

An adequate understanding of young sea turtle dispersal patternsis necessary for effective management of threatened or endangeredspecies. Such patterns are poorly understood, and the term "lostyear" has been adopted to emphasize this gap in sea turtle lifehistory information. Tag returns from pen-reared yearling seaturtles indicate ocean current dispersal. Evidence indicateshatchlings would be similarly dispersed by ocean currents. Feedingstudies with tank-held animals suggest that food resources areavailable in ocean currents for long-term sea turtle survival.Green turtle (Chelonia mydas) growth appears slow in nature.     

Molecular evolution of color vision of zebra finch

We have isolated and sequenced the RH1(Tg), RH2(Tg), SWS2(Tg), and LWS(Tg) opsin cDNAs from zebra finch retinas. Upon binding to 11-cis-retinal, these opsins regenerate the corresponding photosensitive molecules, visual pigments. The absorption spectra of visual pigments have a broad bell shape, with the peak being called lambda(max). Previously, SWS1(Tg) opsin cDNA was isolated from zebra finch retinal RNA, expressed in cultured COS1 cells, reconstituted with 11-cis-retinal, and the lambda(max) of the resulting visual pigment was shown to be 359nm. Here, the lambda(max) values of the RH1(Tg), RH2(Tg), SWS2(Tg), and LWS(Tg) pigments are determined to be 501, 505, 440, and 560nm, respectively. Molecular evolutionary analyses suggest that specific amino acid replacements in the SWS1 and SWS2 pigments, resulting from accelerated evolution, must have been responsible for their functional divergences among the avian pigments.     

Difference in nitrate reduction in "light" and "dark" stages of synchronously grown Chlorella pyrenoidosa and resultant metabolic changes

Using synchronized cells of Chlorella pyrenoidosa, the incorporationpatterns of ¹⁴C into various metabolites with and without nitrogensources were studied under steady-state and non steady-stateconditions. From the patterns it was found that the smallestcells which are divided in the dark utilize nitrate and nitritevery little, if at all. The importance of ammonia for regulation of secondary flow forChlorella is discussed and the suggested regulatory points aredescribed. ¹This work was sponsored, in part, by the U.S. Atomic EnergyCommission (Received January 26, 1970; )     

Behavioral Thermoregulation and the "Final Preferendum" Paradigm

Wider attention to Fry's (1947) "final preferendum" paradigmwould facilitate comparative studies of temperature preference(behavioral thermoregulation) among different animal groups.According to Fry's bipartite definition, the final preferendumis that temperature at which preference and acclimation areequal, and to which an animal in a thermal gradient will finallygravitate regardless of its prior thermal experience (acclimation).This paradigm is helpful in distinguishing between acute thermalpreferenda (measured within 2 hr or less after placing an animalin a thermal gradient), which are influenced by acclimationtemperature, and the species-specific final preferendum (measured24–96 hr after placement in the gradient), which is essentiallyindependent of prior acclimation because reacclimation occursduring the gravitation process. The paradigm does not take intoaccount non-thermal acclimatization influences (e.g., season,photoperiod, age, light intensity, salinity, disease, nutrition,pollutants, biotic interactions) which can also affect temperaturepreference. However, a graph of acutely preferred temperaturesversus acclimation temperatures can be employed to determinean equivalent acclimation temperature for any given acclimatizationstate, as a simple means of quantifying acclimatization statesresulting from interactions of many influences. This paradigm,developed for use with fishes, can also be applied to otherectothermic taxa, although it is most easily employed with aquaticorganisms because of the simplicity of specifying aquatic thermalenvironments in terms of water temperature alone. Methodologiesused in studies of behavioral thermoregulation should take theparadigm into account (especially with respect to length oftests) to enhance the comparative value of data across taxa.     

Juvenile Hormone: The Status of "Status Quo"

SYNOPSIS. Cuticular proteins show specificity for stage, age,and anatomical region. Analysis of the cuticular proteins ofsecond pupae created by application of juvenile hormone demonstratesthat the hormone prevents the onset of new sequences of synthesesand favors repetition of the region-specific, temporal patternof syntheses used in the previous stage. The argument made isthat juvenile hormone might exert this "status quo" action bypreventing alterations in chromatin configuration. Evidencefrom a wide variety of systems shows that polyamines might beinvolved in reprogramming chromatin. Ecdysterone induces ornithinedecarboxylase, the rate limiting enzyme in animal polyaminesynthesis. I suggest that juvenile hormone might be exertingits status quo action by inhibiting this induction. Preliminarystudies of ornithine decarboxylase induction support this specifichypothesis; experiments with an inhibitor of this enzyme, -difluoromethylornithine, however, do not show the expected juvenile hormonemimicry. Further studies are needed to define the control ofpolyamine biosynthesis in insects and to discover whether juvenilehormone plays a role in this control.     

Intronic gene conversion in the evolution of human X-linked color vision genes

Human red and green visual pigment genes are X-linked duplicate genes. To study their evolutionary history, introns 2 and 4 (1,987 and 1,552 bp, respectively) of human red and green pigment genes were sequenced. Surprisingly, we found that intron 4 sequences of these two genes are identical and that the intron 2 sequences differ by only 0.3%. The low divergences are unexpected because the duplication event producing the two genes is believed to have occurred before the separation of the human and Old World monkey (OWM) lineages. Indeed, the divergences in the two introns are significantly lower than both the synonymous divergence (3.2% +/- 1.1%) and the nonsynonymous divergence (2.0% +/- 0.5%) in the coding sequences (exons 1-6). A comparison of partial sequences of exons 4 and 5 of human and OWM red and green pigment genes supports the hypothesis that the gene duplication occurred before the human-OWM split. In conclusion, the high similarities in the two intron sequences might be due to very recent gene conversion, probably during evolution of the human lineage.      

Development of Sink Capacity of the "Storage Root" in a Radish Variety with a Low Ratio of "Storage Root" to Shoot

Controling mechanisms of sink capacity are poorly understood.Previously we suggested that sucrose synthase (SuSy), but notinvertase, plays an important role for sink capacity of theradish "storage root" in a variety, Raphanus sativus L. (cv.White Cherish) [plant Cell Physiol. (1999) 40: 369]. With thisvariety about 50% of the total dry weight (DW) was in the "storageroot" at 21 d after sowing (DAS). We investigated the sink capacityof another radish variety, R. sativus L. (cv. Kosena) with alow ratio of "storage root" to shoot. With the latter varietyonly 3% of the total DW was in the "storage root" at 21 DAS.Sink activity (increase in DW of the "storage root" per unitof DW present per unit of time) of the "storage root" in Kosenaas well as White Cherish was strongly related to the level andactivity of SuSy but not to the activity of invertase. Theseresults confirmed that SuSy rather than invertase may be criticalfor the development of the sink activity of the radish "storageroot" and that the reaction products of UDP-glucose and fructoseare utilized for sink growth including biosynthesis of the cellwall. In Kosena photosynthates seemed to be partitioned mainlyinto developing leaves and fibrous roots. Differences in partitioningof photosynthates among various sinks with these two varietiesare discussed including anatomical considerations. (Received July 19, 1999; Accepted September 30, 1999)     

Development of Sink Capacity of the "Storage Root" in a Radish Cultivar with a High Ratio of "Storage Root" to Shoot

The mechanisms that control sink capacity are poorly understood.in radish, a major sink is the "storage root", which beginsto thicken early in development, mainly as a result of thickeningof the hypocotyl. We investigated changes in the accumulationof dry matter, sink activity (increase in dry weight of thehypocotyl per unit of dry weight present per unit of time),carbohydrate content, levels of metabolites, activities of enzymesrelated to the breakdown of sucrose, and the profile of solubleproteins, as well as changes in anatomy, using hypocotyls ofa cultivar with a high ratio of "storage root" to shoot. Wefound that sink activity was strongly related to the level andactivity of sucrose synthase but not to the activity of invertase.We also found a significant correlation between sucrose contentand the level and activity of sucrose synthase. Our resultssuggest that sucrose synthase, but not invertase, might be criticalfor the development of the sink activity of the radish hypocotyland that the level of sucrose might regulate the expressionof sucrose synthase. A discussion of sink capacity is presentedthat includes consideration of structural changes in the hypocotyl. (Received December 14, 1998; Accepted January 27, 1999)     

Acetate metabolism in the process of "acetate-bleaching" of Chlorella protothecoides

Green cells of Chlorella protothecoides when incubated in amedium containing acetate but no nitrogen source, have beenshown to be bleached as strongly as in glucose-induced bleaching.Using U-¹⁴C-acetate as tracer, the acetate metabolism of algalcells during the process of acetate-induced bleaching was investigated.Changes in algal cell activities for respiration and assimilationof added ¹⁴C-acetate were followed during bleaching processesin "acetate-adapted" and "non-adapted" green cells. As in glucose-inducedbleaching of algal cells, algal cell activity for incorporating¹⁴C into lipids showed the most characteristic change, suggestingthat lipogenesis is causally related to the occurrence of bleachingin algal cells. (Received March 5, 1969; )     

The Essential Role of "Minor" Phyla in Molecular Studies of Animal Evolution

SYNOPSIS. Molecular studies have revealed many new hypothesesof metazoan evolution in recent years. Previously, using morphologicalmethods, it was difficult to relate "minor" animal groups representingmicroscopic metazoans to larger, more well known groups suchas arthropods, molluscs, and annelids. Molecular studies suggestthat acanthocephalans evolved from rotifers, that priapulidsshare common ancestry with all other molting animals (Ecdysozoa),and that flatworms, gnathostomulids and rotifers form a sistergroup to the remaining non-molting protostomes (Lophotrochozoa),together forming Spiralia. The lophophorate phyla (phoronids,brachiopods and bryozoans) appear as protostomes, allied withannelids and molluscs rather than with deuterostomes. Thesefindings present a very different view of metazoan evolution,and clearly show that small and simple animals do not necessarilyrepresent ancestral or primitive taxa.     

The effect of growth regulators on the growth and pigmentation of "Baccara" rose flowers

"Baccara" rose buds were treated with various growth regulatorsduring late stages of bud development. The effect of these substanceson growth and pigmentation were determined. Growth regulatorswere applied by spray or injection or as a lanolin paste, alsoin the nutrient media on which petals were cultured in vitro.Injection of GA into the base of the receptacle caused elongationof the bud whereas IAA, K, ABA, AMO-1618, CCC, and SADH hadlittle or no effect. CCC and MeCl-F did not reduce the elongationcaused by GA. GA treatments also enhanced flower weight andpetal pigmentation and MeCl-F decreased the gibberellin effecton pigmentation. GA treatments of intact flowers and excisedpetals cultured in vitro, were only effective at low temperatures. Gibberellin treatments increased the size of petals, the receptacleand the pedicel only if applied directly to the receptacle.Treatments at lower positions on the flowering shoot eitherhad no effect at all, or caused elongation of only the receptacle. Endogenous gibberellin levels are higher in the receptacle thanin petals or in the pedicel. Injection of GA into the receptaclesignificantly increased gibberellin activity in all flower partswhereas injection into the flowering-shoot base increased gibberellinactivity only in the receptacle. The possibility is discussed that GA, which is exogenously supplieddirectly to the receptacle, enhances flower dimensions and pigmentationby drawing photosynthates to the flower as a consequence ofintensification of the sink. (Received August 17, 1973; )