Genomic evidence for rod monochromacy in sloths and armadillos suggests early subterranean history for Xenarthra

Rod monochromacy is a rare condition in vertebrates characterized by the absence of cone photoreceptor cells. The resulting phenotype is colourblindness and low acuity vision in dim-light and blindness in bright-light conditions. Early reports of xenarthrans (armadillos, sloths and anteaters) suggest that they are rod monochromats, but this has not been tested with genomic data. We searched the genomes of Dasypus novemcinctus (nine-banded armadillo), Choloepus hoffmanni (Hoffmann''s two-toed sloth) and Mylodon darwinii (extinct ground sloth) for retinal photoreceptor genes and examined them for inactivating mutations. We performed PCR and Sanger sequencing on cone phototransduction genes of 10 additional xenarthrans to test for shared inactivating mutations and estimated the timing of inactivation for photoreceptor pseudogenes. We concluded that a stem xenarthran became an long-wavelength sensitive-cone monochromat following a missense mutation at a critical residue in SWS1, and a stem cingulate (armadillos, glyptodonts and pampatheres) and stem pilosan (sloths and anteaters) independently acquired rod monochromacy early in their evolutionary history following the inactivation of LWS and PDE6C, respectively. We hypothesize that rod monochromacy in armadillos and pilosans evolved as an adaptation to a subterranean habitat in the early history of Xenarthra. The presence of rod monochromacy has major implications for understanding xenarthran behavioural ecology and evolution.     

Spectral tuning by opsin coexpression in retinal regions that view different parts of the visual field

Vision frequently mediates critical behaviours, and photoreceptors must respond to the light available to accomplish these tasks. Most photoreceptors are thought to contain a single visual pigment, an opsin protein bound to a chromophore, which together determine spectral sensitivity. Mechanisms of spectral tuning include altering the opsin, changing the chromophore and incorporating pre-receptor filtering. A few exceptions to the use of a single visual pigment have been documented in which a single mature photoreceptor coexpresses opsins that form spectrally distinct visual pigments, and in these exceptions the functional significance of coexpression is unclear. Here we document for the first time photoreceptors coexpressing spectrally distinct opsin genes in a manner that tunes sensitivity to the light environment. Photoreceptors of the cichlid fish, Metriaclima zebra, mix different pairs of opsins in retinal regions that view distinct backgrounds. The mixing of visual pigments increases absorbance of the corresponding background, potentially aiding the detection of dark objects. Thus, opsin coexpression may be a novel mechanism of spectral tuning that could be useful for detecting prey, predators and mates. However, our calculations show that coexpression of some opsins can hinder colour discrimination, creating a trade-off between visual functions.     

Molecular basis of spectral tuning in the newt short wavelength sensitive visual pigment

Previously we reported the sequence of the member of the short wavelength sensitive 2 (SWS2) family of vertebrate visual pigments from the retina of the Japanese common newt, Cynops pyrrhogaster[Takahashi, Y. et al. (2001) FEBS Lett. 501, 151-155]. Now we have expressed the apopigment and regenerated it with A1 retinal. Its absorption maximum, 474 nm, is greatly red shifted compared to other known SWS2 pigments (418-455 nm). To determine the amino acid residues that control its spectral tuning, we replaced the residues that were near the chromophore and which differed between the newt and the bullfrog (lambda(max) = 430 nm) wild-type SWS2 pigments: Pro91Ser, Ser94Ala, Ile122Met, Cys127Ser, Ser211Cys, Tyr261Phe, and Ala292Ser. Each of these site-directed mutants led to blue shifts of the newt pigment with five of them causing substantial shifts; their sum was about equal to the difference between the absorption maximum of the bullfrog and newt pigments, 44 nm. The 32 nm shift of the absorption maximum of the multiple seven-residue mutant to 442 nm is fairly close to that of the wild-type bullfrog pigment. Thus, the seven amino acid residues that we replaced are the major cause of the red shift of the newt SWS2 pigment's spectrum. Two of the residues, 91 and 94, have not previously been identified as wavelength regulating sites in visual pigments. One of these, 91, probably regulates color via a new mechanism: altering of a hydrogen bonding network that is connected via a water to the chromophore, in this case its counterion, Glu113.     

A novel amino acid substitution is responsible for spectral tuning in a rodent violet-sensitive visual pigment

Cone short-wave (SWS1) visual pigments can be divided into two categories that correlate with spectral sensitivity, violet sensitive above 390 nm and ultraviolet sensitive below that wavelength. The evolution and mechanism of spectral tuning of SWS1 opsins are proving more complex than those of other opsin classes. Violet-sensitive pigments probably evolved from an ancestral ultraviolet-sensitive opsin, although in birds ultraviolet sensitivity has re-evolved from violet-sensitive pigments. In certain mammals, a single substitution involving the gain of a polar residue can switch sensitivity from ultraviolet to violet sensitivity, but where such a change is not involved, several substitutions may be required to effect the switch. The guinea pig, Cavia porcellus, is a hystricognathous rodent, a distinct suborder from the Sciurognathi, such as rats and mice. It has been shown by microspectrophotometry to have two cone visual pigments at 530 and 400 nm. We have ascertained the sequence of the short-wave pigment and confirmed its violet sensitivity by expression and reconstitution of the pigment in vitro. Moreover, we have shown by site-directed mutagenesis that a single residue is responsible for wavelength tuning of spectral sensitivity, a Val86Phe causing a 60 nm short-wave shift into the ultraviolet and a Val86Tyr substitution shifting the pigment 8 nm long wave. The convergent evolution of this mammalian VS pigment provides insight into the mechanism of tuning between the violet and UV.     

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.     

Evolution and mechanism of spectral tuning of blue-absorbing visual pigments in butterflies

The eyes of flower-visiting butterflies are often spectrally highly complex with multiple opsin genes generated by gene duplication, providing an interesting system for a comparative study of color vision. The Small White butterfly, Pieris rapae, has duplicated blue opsins, PrB and PrV, which are expressed in the blue (λ _max = 453 nm) and violet receptors (λ _max = 425 nm), respectively. To reveal accurate absorption profiles and the molecular basis of the spectral tuning of these visual pigments, we successfully modified our honeybee opsin expression system based on HEK293s cells, and expressed PrB and PrV, the first lepidopteran opsins ever expressed in cultured cells. We reconstituted the expressed visual pigments in vitro, and analysed them spectroscopically. Both reconstituted visual pigments had two photointerconvertible states, rhodopsin and metarhodopsin, with absorption peak wavelengths 450 nm and 485 nm for PrB and 420 nm and 482 nm for PrV. We furthermore introduced site-directed mutations to the opsins and found that two amino acid substitutions, at positions 116 and 177, were crucial for the spectral tuning. This tuning mechanism appears to be specific for invertebrates and is partially shared by other pierid and lycaenid butterfly species.     

A genetic assessment of polyandry and breeding-site fidelity in lemon sharks

We here employ 11 microsatellite markers and recently developed litter reconstruction methods to infer mating system parameters (i.e. polyandry and breeding-site fidelity) at a lemon shark nursery site in Marquesas Key, Florida. Four hundred and eight juvenile or subadult sharks were genotyped over eight complete breeding seasons. Using this information, we were able to infer family structure, as well as fully or partially reconstruct genotypes of 46 mothers and 163 fathers. Multiple litter reconstruction methods were used, and novel simulations helped define apparent bias and precision of at least some mating system parameters. For Marquesas Key, we find that adult female lemon sharks display high levels of polyandry (81% of all litters sampled) and stronger fidelity to the nursery site than do males. Indeed, few male sharks sired offspring from more than one litter during the course of the study. These findings were quite similar to previous results from another lemon shark nursery site (Bimini, Bahamas), suggesting conserved mating system parameters despite significant variation in early life-history traits (i.e. body size and growth) among sites. The finding of at least some site fidelity in females also supports the need for careful conservation of each nursery.     

Polymorphic microsatellite markers for studies of the conservation and reproductive genetics of imperilled sand tiger sharks (Carcharias taurus)

We report on the isolation of eight microsatellites from the sand tiger shark, Carcharias taurus, using an enrichment protocol. All loci, with the exception of Cta45–183, were in Hardy–Weinberg equilibrium. Loci exhibited three to 15 alleles, and observed and expected heterozygosities of 0.095–1.000 and 0.284–0.924, respectively. An additional marker (Iox‐12) developed from a shortfin mako library was variable in sand tigers. These markers will be used to examine population genetics and mating patterns of this imperilled species.     

On visual pigment templates and the spectral shape of invertebrate rhodopsins and metarhodopsins

The absorbance spectra of visual pigments can be approximated with mathematical expressions using as single parameter the absorbance peak wavelength. A comparison of the formulae of Stavenga et al. in Vision Res 33:1011–1017 (1993) and Govardovskii et al. in Vis Neurosci 17:509–528 (2000) applied to a number of invertebrate rhodopsins reveals that both templates well describe the normalized α-band of rhodopsins with peak wavelength > 400 nm; the template spectra are virtually indistinguishable in an absorbance range of about three log units. The template formulae of Govardovskii et al. in Vis Neurosci 17:509–528 (2000) describe the rhodopsin spectra better for absorbances below 10⁻³. The template predicted spectra deviate in the ultraviolet wavelength range from each other as well as from measured spectra, preventing a definite conclusion about the spectral shape in the wavelength range <400 nm. The metarhodopsin spectra of blowfly and fruitfly R1-6 photoreceptors derived from measured data appear to be virtually identical. The established templates describe the spectral shape of fly metarhodopsin reasonably well. However, the best fitting template spectrum slightly deviates from the experimental spectra near the peak and in the long-wavelength tail. Improved formulae for fitting the fly metarhodopsin spectra are proposed.     

Diversification and spectral tuning in marine proteorhodopsins

Proteorhodopsins, ubiquitous retinylidene photoactive proton pumps, were recently discovered in the cosmopolitan uncultured SAR86 bacterial group in oceanic surface waters. Two related proteorhodopsin families were found that absorb light with different absorption maxima, 525 nm (green) and 490 nm (blue), and their distribution was shown to be stratified with depth. Using structural modeling comparisons and mutagenesis, we report here on a single amino acid residue at position 105 that functions as a spectral tuning switch and accounts for most of the spectral difference between the two pigment families. Furthermore, looking at natural environments, we found novel proteorhodopsin gene clusters spanning the range of 540-505 nm and containing changes in the same identified key switch residue leading to changes in their absorption maxima. The results suggest a simultaneous diversification of green proteorhodopsin and the new key switch variant pigments. Our observations demonstrate that this single-residue switch mechanism is the major determinant of proteorhodopsin wavelength regulation in natural marine environments.     

Map-like use of Earth’s magnetic field in sharks

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The importance of habitat and life history to extinction risk in sharks, skates, rays and chimaeras

We compared life-history traits and extinction risk of chondrichthyans (sharks, rays and chimaeras), a group of high conservation concern, from the three major marine habitats (continental shelves, open ocean and deep sea), controlling for phylogenetic correlation. Deep-water chondrichthyans had a higher age at maturity and longevity, and a lower growth completion rate than shallow-water species. The average fishing mortality needed to drive a deep-water chondrichthyan species to extinction (Fextinct) was 38-58% of that estimated for oceanic and continental shelf species, respectively. Mean values of Fextinct were 0.149, 0.250 and 0.368 for deep-water, oceanic and continental shelf species, respectively. Reproductive mode was an important determinant of extinction risk, while body size had a weak effect on extinction risk. As extinction risk was highly correlated with phylogeny, the loss of species will be accompanied by a loss of phylogenetic diversity. Conservation priority should not be restricted to large species, as is usually suggested, since many small species, like those inhabiting the deep ocean, are also highly vulnerable to extinction. Fishing mortality of deep-water chondrichthyans already exploited should be minimized, and new deep-water fisheries affecting chondrichthyans should be prevented.     

Molecular ecology and adaptation of visual photopigments in craniates

In craniates, opsin‐based photopigments expressed in the eye encode molecular ‘light sensors’ that constitute the initial protein in photoreception and the activation of the phototransduction cascade. Since the cloning and sequencing of the first vertebrate opsin gene (bovine rod opsin) nearly 30 years ago (Ovchinnikov Yu 1982, FEBS Letters, 148, 179–191; Hargrave et al. 1983, Biophysics of Structure & Mechanism, 9, 235–244; Nathans & Hogness 1983, Cell, 34, 807–814), it is now well established that variation in the subtypes and spectral properties of the visual pigments that mediate colour and dim‐light vision is a prevalent mechanism for the molecular adaptation to diverse light environments. In this review, we discuss the origins and spectral tuning of photopigments that first arose in the agnathans to sample light within the ancient aquatic landscape of the Early Cambrian, detailing the molecular changes that subsequently occurred in each of the opsin classes independently within the main branches of extant jawed gnathostomes. Specifically, we discuss the adaptive changes that have occurred in the photoreceptors of craniates as they met the ecological challenges to survive in quite differing photic niches, including brightly lit aquatic surroundings; the deep sea; the transition to and from land; diurnal, crepuscular and nocturnal environments; and light‐restricted fossorial settings. The review ends with a discussion of the limitations inherent to the ‘nocturnal‐bottleneck’ hypothesis relevant to the evolution of the mammalian visual system and a proposition that transition through a ‘mesopic‐bottleneck’ may be a more appropriate model.     

Introduction of hydroxyl-bearing amino acids causes bathochromic spectral shifts in rhodopsin. Amino acid substitutions responsible for red-green color pigment spectral tuning.

Comparisons of the deduced amino acid sequences of eight primate photopigment genes led to the proposal that three amino acid substitutions produce the approximately 1,000 cm-1 difference in the absorption maxima of human red and green pigments (Neitz, M., Neitz, J., and Jacobs, G.H. (1991) Science 252, 971-974). We tested this proposal by mutating these three residues in rhodopsin and evaluating the effects on spectral properties. Nonpolar residues normally present in rhodopsin and in the green pigment were substituted by hydroxyl-bearing residues normally present in the red pigment. Two of these substitutions (Phe-261 to Tyr or Ala-269 to Thr) caused significant red shifts in the absorption maxima of the resulting mutant pigments. A third substitution (Ala-164 to Ser) caused only a slight effect. Combinations of substitutions caused additive shifts in absorption maxima. A double mutant (Phe-261 to Tyr/Ala-269 to Thr) displayed an absorption maximum that was red-shifted by 775 cm-1. Wavelength modulation in the visual pigments responsible for red-green color vision is likely to be governed by retinal-protein interactions involving primarily these two amino acid residues. Furthermore, interactions of hydroxyl-bearing amino acids with the chromophore may be a general mechanism of the opsin shift in visual pigments.     

柏科4个属(Fokienia、Cupressus、Chamaecyparis和Juniperus)植物雌球果的发育(英文)

通过扫描电镜和常规石蜡切片技术,观察了柏科4个属(Fokienia、Cupressus、Chamaecyparis和Juniperus)植物雌球果中胚珠的发育及苞片的结构变化。在所有研究的种类中,可育苞片腋部最先观察到的结构是一扁平的突起,并在其上分化出发育为胚珠的胚珠原基。在雌球果的发育过程中,未观察到独立的珠鳞发育。不同的种中,胚珠的数量和发育顺序有所不同,但苞片的发育是相似的。传粉前,苞片的结构与叶相似。传粉后,由于剧烈的居间生长,苞片发育为盾形,形成球形的球果。另外,在发育后期,苞片内维管系统变得复杂,并且在近轴面有反向的维管束发育。我们还对柏科植物雌球果的形态学特性及其可能的演化趋势进行了讨论。     

Effect of ration size on growth and gross conversion efficiency of young lemon sharks, Negaprion brevirostris

Young lemon sharks, Negaprion brevirostris , were kept under controlled conditions in an aquarium and fed blue runner, Caranx crysos , at different ration levels. The relationship between feeding rate and growth rate was best described by a von Bertalanffy growth curve, which predicted a maximum growth rate of 140 kJ kg⁻¹ day⁻¹ (0·66% b.w. day⁻¹), a maintenance ration of 199 kJ kg⁻¹ day⁻¹ (1·06% b.w. day⁻¹), and losses due to starvation of -236kJ kg⁻¹ day⁻¹ (1·11% b.w. day⁻¹). The relationship between gross conversion efficiency ( K ₁) and feeding rate was also examined. K₁ ranged from - 64 to 25% and did not drop at high ration levels. Activity levels of both starved sharks and sharks fed at maintenance were not significantly different (0·2 body lengths s⁻¹). K ₁ values generated from both laboratory and field data suggest that young lemon sharks can convert food to new tissue as efficiently as teleosts.