Major Vertebrate Classes Research Articles

Fourth component of Xenopus laevis complement: cDNA cloning and linkage analysis of the frog MHC.

Complement C4 shows extensive structural and functional similarity to complement C3, hence these components are believed to have originated by gene duplication from a common ancestor. Although to date C3 cDNA clones have been isolated from all major classes of extant vertebrates including Xenopus, C4 cDNA clones have been isolated from mammalian species only. We describe here the molecular cloning and structural analysis of Xenopus C4 cDNA. The cDNA sequence encoding the thioester region of Xenopus C4 was amplified by reverse transcriptase-polymerase chain reaction using Xenopus liver mRNA as a template, and then used to screen a liver cDNA library. The amino acid sequence of Xenopus C4 deduced from a clone containing the entire protein-coding sequence showed 39%, 30%, 25%, and 20% overall identity with those of human C4, C3, C5, and alpha2-macroglobulin, respectively. The predicted amino acid sequence consisted of a 22-residue putative signal peptide, a 634-residue beta chain, a 732-residue alpha chain, and a 287-residue gamma chain. Of 30 cysteine residues, 27 were found in exactly the same positions as in human C4. Genomic Southern blotting analysis indicated that C4 is a single copy gene in Xenopus and is part of the frog MHC cluster. These results clearly demonstrate that C3/C4 gene duplication and linkage between the C4 gene and the major histocompatibility complex predate mammalian/amphibian divergence.

Three retroviral sequences in amphibians are distinct from those in mammals and birds

We isolated and characterized three endogenous retroviral fragments from the dart-poison frog Dendrobates ventrimaculatus. These are the first retroviral sequences to be identified in amphibians, and consequently retroviruses have now been found in each of the five major vertebrate classes. Comparison of the amphibian retroviral fragments, termed DevI, DevII, and DevIII, with mammalian and avian isolates revealed significant differences between their nucleotide sequences. This suggested that they were only distantly related to the seven currently recognized retroviral genera. Additional analysis by phylogeny reconstruction showed that the amphibian retroviral fragments were approximately equally related to the Moloney leukemia-related viruses, the spumaviruses, and walleye dermal sarcoma virus. Hybridization experiments revealed that viruses closely related to DevI, DevII, and DevIII do not appear to be widespread in other vertebrates and that DevI, DevII, and DevIII are all present at high copy numbers within their amphibian hosts, typically at over 250 copies per genome. The viruses described here, along with two others which have recently been found in a fish and a reptile, indicate that there may be some major differences in the retroviruses harbored by different vertebrate classes. This suggests that further characterization of retroviruses of fish, reptiles, and amphibians will help in understanding the evolution of the whole retroviral family and may well lead to the discovery of retroviruses with novel biological properties.

Comparative distribution of neurotensin‐like immunoreactivity in the brain of a teleost (carassius auratus), an amphibian (hyla meridionalis), and a reptile (gallotia galloti)

The distribution of neurotensin (NT) was studied in the brain of three species belonging to the three major classes of cold-blooded vertebrates: teleost fishes (Carassius auratus), anuran amphibians (Hyla meridionalis), and reptiles (Gallotia galloti; Lacertidae). By using antibodies directed against synthetic bovine NT in the three species, immunoreactive cell bodies were discovered mostly in the telencephalon and diencephalon, in particular at the level of the preoptic region the mediobasal hypothalamus, and the thalamus. In the frog and the lizard, additional immunoreactive (ir) structures were observed in the optic tectum and the tegmentum of the mesencephalon. In the goldfish pituitary, an extensive innervation was consistently observed at the level of the rostral pars distalis, whereas in both frog and lizard, positive fibers were only detected in the external layer of the median eminence. In the three species there is a striking overlap between the distribution of the NT-ir cell bodies and that of the target cells for sexual steroids. The results are discussed in relation with those reported in birds and mammals, and with the possible interactions among NT, sexual steroids, and the neuroendocrine control of pituitary hormone release, in particular prolactin and gonadotrophin.

The complete amino acid sequence of prolactin from the bullfrog, Rana catesbeiana

The complete amino acid sequence of prolactin (PRL) from an amphibian species, the bullfrog ( Rana catesbeiana), has been determined and conserved residues and domains were analyzed by sequence comparison of PRLs from 15 species of five major vertebrate classes. The bullfrog PRL consists of 197 amino acid residues with three disulfide linkages formed between residues 4–11, 58–172, and 189–197. The bullfrog PRL shows the highest identity with sea turtle PRL (75%); lower identities with chicken PRL (72%), pig, horse, and fin whale PRLs (68%), human, cattle, sheep, and elephant PRLs (60-58%), and rat and mouse PRLs (50%); and significantly lower identity with teleost PRLs (about 30%). It is apparent that all tetrapod PRLs characterized so far contain three disulfide bonds in homologous positions and differ from teleost PRLs which lack the N-terminal disulfide loop. The tetrapod and teleost PRLs share 34 common residues and these conserved residues are clustered in six domains (PD1 to PD6), suggesting that these common residues, or at least part of them, are responsible for the activities common to all PRLs. On the other hand, PD5 is conserved significantly within tetrapod PRLs, but to a lesser extent in teleos PRLs, suggesting that the PD5 contributes to the activities specific to tetrapod PRLs.

The protein sequence homology of gamma-crystallins among major vertebrate classes and their DNA sequence homology to heat-shock protein genes.

A systematic characterization of lens crystallins from five major classes of vertebrates was carried out by exclusion gel filtration, cation-exchange chromatography and N-terminal sequence determination. All crystallin fractions except that of gamma-crystallin were found to be N-terminally blocked. gamma-Crystallin is present in major classes of vertebrates except the bird, showing none, or decreased amounts, of this protein in chicken and duck lenses, respectively. N-Terminal sequence analysis of the purified gamma-crystallin polypeptides showed extensive homology between different classes of vertebrates, supporting the close relatedness of this family of crystallin even from the evolutionarily distant species. Comparison of nucleotide sequences and their predicted amino acid sequences between gamma-crystallins of carp and rat lenses and heat-shock proteins demonstrated partial sequence homology of the encoded polypeptides and striking homology at the gene level. The unexpected strong homology of complementary DNA (cDNA) lies in the regions coding for 40 N-terminal residues of carp gamma-II, rat gamma 2-1, and the middle segments of 23,000- and 70,000-Mr heat-shock proteins. The optimal alignment of DNA sequences along these two segments shows about 50% homology. The percentage of protein sequence identity for the corresponding aligned segments is only 20%. The weak sequence homology at the protein level is also found between the invertebrate squid crystallin and rat gamma-crystallin polypeptides. These results pointed to the possibility of unifying three major classes of vertebrate crystallins into one alpha/beta/gamma superfamily and corroborated the previous supposition that the existing crystallins in the animal kingdom are probably mutually interrelated, sharing a common ancestry.

Biochemical comparison of lens crystallins from three reptilian species

Lens crystallins were isolated from the homogenates of reptilian eye lenses derived from three different species by gel-permeation chromatography and chartcterized by gel electrophoresis, amino-acid analysis, N-terminal sequence analysis and circular dichroism. Four fractions corresponding to α-, δ / ε / β- β- and γ-crystallins were obtained for the crystallins from caiman lenses, whereas δ- and γ-crystallin fraction were present in lesser amounts or missing in the turtle and snake lenses, respectively. The native molecular masses for these purified fractions and their polypeptide compositions were determined by gel filtration and SDS-gel electrophoresis, respectively, revealing the typical subunit compositions for each classified crystallin. The spectra of circular dichroism indicate a predominant β-sheet structure in α-, β- and γ-crystallins, and a major contribution of α-helical structure in δ / ε-crystallin fraction, which bears a resemblance to the secondary structure of δ-crystallin from the chicken lenses. Comparison of the amino-acid contents of each orthologous class of reptillan crystallins with those of evolutionary distant species still exhibited similarity in their amino-acid compositions. N-terminal sequence analysis of the crystallin fractions revealed that all fractions except that of γ-crystallin are N-terminally blocked. Extensive sequence similarity between the reptilian γ-crystallin polypeptides and those from other vertebrate species were found, which establish the close relatedness of γ-crystallins amongst the major classes of vertebrates.

Antigenic sites common to major fish myelin glycoproteins (IP) and to major tetrapod PNS myelin glycoprotein (Po) reside in the amino acid chains

The major myelin glycoproteins in the CNS and PNS of trout (IP) were enzymatically deglycosylated with endoglycosidase F (Endo F) and examined by electro-immunoblotting. Following carbohydrate removal and loss of concanavalin A affinity each of the four IP components underwent a similar reduction in molecular size, corresponding to approximately 3,000 daltons. Immunological cross-reactivities with anti-bovine Po or anti-trout IP2 antibodies, were however fully retained by the Endo F cleavage products. This strongly implies that the antigenic sites shared by the mammalian Po protein and the various intermediate glycoproteins of trout CNS and PNS are located in the protein portion. Immunoblot analysis of the PNS myelin proteins from various species of the major vertebrate classes with anti-trout IP2 antiserum revealed striking differences in the immunological properties of the individual Po components which were not detected when anti-bovine Po antiserum was used as a probe.

Phylogenetic comparison of lens crystallins from the vertebrate and invertebrate - convergent or divergent evolution?

A systematic biochemical comparison has been made of the crystallins isolated from the lenses of five different species belonging to the five major classes of vertebrates. Gel-permeation chromatography of the lens homogenates on Fractogel TSK HW-55(S) revealed well-defined elution patterns with a characteristic distribution of different classes of crystallins from each species. SDS gel electrophoresis and statistical comparison of the amino acid contents indicated that all crystallin groups from different classes share some common subunits and similarity in their amino acid compositions. The results coupled with the relatedness shown in the amino acid compositions of fish γ-crystallin with those of mammalian γ-crystallin and the squid crystallin from the invertebrate pointed to the possibility of the existence of a common ancestral protein for all crystallins. This is in favor of the divergent rather than convergent evolution of lens crystallins as commonly assumed in the literature.

Functional Anatomy and Evolution of Monoaminergic Systems

1. The organization of brain neuronal systems utilizing monoamines (primary techolamines and serotonin) as neurotransmitters and/or neuromodulators, has been studied in representatives of each major vertebrate class using fluorescence histochemical methods. This long-term investigation has led to the following conclusions: 2. (1) The monoaminergic neurons seem to have occurred early in phylogeny. These neurons, particularly the serotoninergic systems, are already well-developed in the brains of cyclostomes (lampreys) and primitive bony fishes. 3. (2) The monoaminergic cell bodies are especially abundant within the hypothalamus in anamniotes whereas they predominate at brainstem levels in amniotes. However, a similar pattern of distribution of monoaminergic cell bodies has been found in the brainstem of all vertebrates. 4. (3) The monoaminergic neurons may act as an important informative link between the cerebrospinal fluid and the neural tissue in all vertebrates through processes contacting the ventricular space. 5. (4) The monoamingeric neuronal systems are widespread and highly collateralized inall vertebrates. A single brainstem monoaminergic neuron may send divergent axon collaterals to various remote forebrain areas and, thus, could strongly influence vast neuronalassemblies. 6. (5) The terminal arborization of monoaminergic systems, particularly that of the serotoninergicneurons, displays patterns that are strikingly constant from one vertebrate class to the other. Such a constancy strongly suggests that these chemospecific neuronal systems are indeedphylogenetically ancient and subserve similar fundamental brain functions, such as the controlof sleep-waking cycle and of feeding and reproductive behaviors, in all vertebrates.

Mammalian Diversity and Structure

Mammalia constitute one of the five major classes of vertebrates (the others being Pisces, Amphibia, Reptilia, and Aves). There are, at latest count, some 4,170 species of mammals living today, classified in about 1010 genera, 130–140 families, and 18 orders (Corbet and Hill, 1980; Honacki et al., 1982). In addition, sixteen orders and uncounted genera and species of mammals are known to be extinct (see synoptic classification in Table 1). While most living mammals are terrestrial like their Mesozoic ancestors, one large progressive group has invaded the air (bats, represented by about 150 living genera and 920 species). Two other smaller groups are fully aquatic (whales, including some 40 genera and 80 species; and sirenians, 3 genera and 5 species). Mammals occupy all of the continents, and all of the larger oceanic islands. Faunal diversity is greatest in lowland tropical forests, woodlands, and savannas, decreasing rapidly at high elevations and latitudes.

Animal cultures and a general theory of cultural evolution

A review of evidence reveals that animal traditions are phylogenetically widespread, appearing in all major vertebrate classes and in the Insecta. Some, but not all, of these traditions fit a general definition of culture, and the evidence further indicates that the cultural traditions of some avian and mammalian species may constitute complex cultural systems. The general definition of culture employed here is based on a concensus of anthropological thought and on population thinking. Population thinking is then used to develop a general theory of branching-reticulate cultural evolution. This theory identifies the causal mechanisms for change and the resultant patterns of change in human and nonhuman cultures. Cultural evolution is viewed at three levels: microevolution; evolution of cultural institutions; and the evolution of cultural lineages and systems. The parallels with, and differences between, organic and cultural evolution are discussed in detail. The theory of branching-reticulate cultural evolution is then applied to specific instances of change in several avian and mammalian cultures.

Immunochemical and biological studies with antiserum to shark growth hormone

A comparative immunochemical study was conducted with an antiserum prepared for the first time against growth hormone (GH) from a phylogenetically ancient fish, the blue shark ( Prionace glauca). The result of immunodiffusion studies revealed extensive immunochemical relatedness of GHs from species representing each of the major vertebrate classes. The findings obtained with a homologous radioimmunoassay (RIA) system did not show good correlation with those obtained by immunodiffusion. However, a heterologous system employing the same antiserum and a purified rat GH for standards and for the tracer, gave results that were in good agreement with the findings based on immunodiffusion. Interestingly, in the heterologous RIA, the purified GH of a modern bony fish ( Tilapia) as well as the presumptive GH in pituitary extracts of several other modern bony fishes (teleosts), showed significant relatedness to shark GH only when employed at very high concentrations. The rat tibial plate-stimulating activity shown by a shark pituitary extract was completely blocked with injections of antisera to purified shark or turtle GH. Furthermore, purified shark GH showed a very significant stimulation in the rat tibia assay, in contrast to the lack of stimulation reported earlier by several investigators using pituitary extracts of teleostean species and the relatively low degree of stimulation shown recently with a highly purified GH from a teleost ( Tilapia). The data obtained thus far based on immunochemical and biological studies suggest that there must be some significant differences in the structure of shark GH compared to the structure of GH from modern bony fishes, in spite of the existence of substantial similarities.

Kynureninase-type enzymes and the evolution of the aerobic tryptophan-to-nicotinamide adenine dinucleotide pathway

Kynureninase-type ( l-kynurenine hydrolase, EC 3.7.1.3) activity has been found to be present in the livers of fish, amphibia, reptiles, and birds. In addition to past information concerning this enzyme activity in mammalian liver, it is now clear that all the major classes of vertebrates carry a highly specialized kynureninase-type enzyme, which we have termed a hydroxykynureninase. To compare the reactivities of these enzymes with l-kynurenine and l-3-hydroxykynurenine, ratios of τ values ( K m /V ) were used. Based on this comparison, the bacterium Pseudomonas fluorescens carries the most efficient kynureninase, whereas the amphibian Xenopus laevis has the most efficient hydroxykynureniase. In these two cases, the ratio of τ values differs by a factor of 38 000. It is hypothesized that the tryptophan-to-nicotinamide adenine dinucleotide biosynthetic pathway evolved from a catabolic system of enzymes, and that the differences observed in the kynureninase-type enzymes between lower and higher organisms reflect the specialization of the function of these enzymes from a strictly catabolic role to an anabolic one during the course of evolution.

Evolution and Ecological Value of Duplicate Genes

The existence of duplications of parts of chromosomes and of specific genetic loci was inferred in Drosophila by Bridges (39) and Sturtevant (280) early in this cen­ tury. In his address as the retiring president of the American Society of Naturalists in 1947, Metz (201 ), however, could point to only a few specific instances of duplicate bands after cytological studies on the polytene chromosomes from Dipt­ eran insects. Stephens, in a 195 1 review of the significance of duplications in evolu­ tion, concluded that there was not a single proven case in which two gene products could be homologously related (273). These and other students of evolution [see (273) for references] were certainly aware that new genetic material must come from duplications of previously existing genes or chromosomes. Metz even surmised correctly that some duplicate genes might be unstable and that heterochromatin may contain repeated chromosomal parts. Nevertheless, before the advent of molecular biology, essentially only speculation about the extent and the �elective importance of duplicate genes was available. Now, however, while there is still no shortage of speculation about this phenomenon and its evolutionary meaning, there are many firmly documented cases of duplicate genes, including those in which the members of the gene pair are at several different levels of divergence. Kimura & Ohta (172) now list gene duplication as one of five principles of molecular evolu­ tion, and, in addition, Ohno (213) has written a book stressing its importance during the evolution of the major vertebrate classes. This article reviews some of the information from molecular biology which, in the last fifteen years, has effected this change. Soon after the identification of DNA as the genetic material, the extent of the evolutionary increase in the number of genes per organism became evident. The amount of DNA per genome (expressed as picograms or nucleotide pairs) ranges over eight orders of magnitude, from certain simple viruses to some algae, amphibi-

A Comparative Study of Crosslinked and Noncrosslinked Fibrin from the Major Classes of Vertebrates

SummaryCrosslinked and noncrosslinked fibrin formed by clotting whole plasma in the presence and absence of calcium has been examined by polyacrylamide gel electrophoresis in sodium dodecyl sulfate. The fibrin from 45 species of vertebrates with representatives from all classes except the Bradyodonti (Chimaeras) and a majority of the subclasses have been compared. Noncrosslinked fibrin from the mammals contained subunits resembling the α, β, and γ chains of human noncrosslinked fibrin. The molecular weight of the α-chains varied greatly, the molecular weight of the β-chains varied slightly, while the molecular weight of the γ-chains was apparently invariant. Nonerosslinked avian fibrins showed clearly resolved γ-chains with a molecular weight slightly smaller than human γ-chains. Avian α- and β-chains were usually not resolved. Nonerosslinked fibrin from the reptiles and amphibians contained a clearly resolved subunit with a molecular weight similar to that of human γ-chain. In crosslinked fibrin from all of the mammals, birds, reptiles, and amphibians the γ-chain was absent and a new band which corresponded to a dimer of the γ-chain was found, while high molecular weight polymers were only found in a few species. Fibrin formed from the plasma of the bony fishes was often difficult because of the problem of fibrinolysis ; however, non crosslinked fibrin from some species of bony fish had three clearly resolved subunits while the crosslinked fibrin from all of the species examined had dimers as the predominant crosslinked forms. The noncrosslinked fibrin from the cartilaginous fish had subunits in the same molecular weight range as the other vertebrate fibrins; however, the crosslinked fibrin was unique to this class because all of the fibrin subunits were involved in the crosslinking process. Dimers appeared to be a transient species, and a large number of different high molecular weight crosslinked species were formed. The crosslinked fibrin from the hagfish contained dimers and no higher molecular weight crosslinked forms. Rapid fibrinolysis complicated the interpretation of the results from all of the classes of fish.

The Carbohydrate Composition of Immunoglobulins from Diverse Species of Vertebrates

Abstract Immunoglobulins and their respective heavy (H) and light (L) polypeptide chains from species representing the major classes of vertebrates were analyzed for their carbohydrate composition by gas chromatography of the alditol acetate derivatives of the monosaccharides released by acid hydrolysis. Mannose, galactose, glucosamine and sialic acid were present in the immunoglobulins from all the species investigated. Fucose however was absent from the immunoglobulins of two species of animals studied. Most of the carbohydrate was found associated with the H chains. The carbohydrate composition was felt to be heterogeneous in that the quantities of fucose and sialic acid found often accounted for less than one residue per H chain. Whereas species representative of the mammals and birds had mannose to galactose ratios greater than one, the ratio of these sugars to each other in the immunoglobulins from animals below the birds was on the order of one.

Isolation and characterization of immunoglobulin-like proteins of the Australian lungfish (Neoceratodus forsteri).

SummaryProteins resembling immunoglobulins of other vertebrate species in physical and chemical properties were instigated from the serum of unimmunized Australian lungfish (Neoceratodus forsteri). Two types of antigenically related proteins, characterized by sedimentation coefficients of 19.4S and 5.9S, were observed. Both proteins possessed light polypeptide chains which resembled typical immunoglobulin light chains in molecular weight (23,000) and behaviour in gel electrophoresis. The 19.4S proteins were similar to γM immunoglobulins of other vertebrates in size and chain structure. Heavy polypeptide chains from these molecules were comparable to the μ‐chain in molecular weight (70,000) and gel electrophoretic properties. The heavy chain of the 5.9S immunoglobulin‐like protein differed from those of the major classes of vertebrates in possessing a molecular weight of approximately 40,000. In both proteins the light chains and heavy chains were linked through disulfide bonds.These data suggest that lungfish resemble higher vertebrates in possessing different immunoglobulin classes defined by the presence of distinct heavy chains. The low molecular weight type of immunoglobulin‐like proteins may represent a class of immunoglobulins which is unique to Dipnoi.