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Jacek M Szymura - One of the best experts on this subject based on the ideXlab platform.
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a dense linkage map for a large repetitive genome discovery of the sex determining region in hybridising fire bellied toads Bombina Bombina and b variegata
bioRxiv, 2020Co-Authors: Beate Nurnberger, Mark Blaxter, S J E Baird, D Cizkova, A Bryjova, Austin B Mudd, Jacek M SzymuraAbstract:Abstract Hybrid zones that result from secondary contact between diverged populations offer unparalleled insight into the genetic architecture of emerging reproductive barriers and so shed light on the process of speciation. Natural selection and recombination jointly determine their dynamics, leading to a range of outcomes from finely fragmented mixtures of the parental genomes that facilitate introgression to a situation where strong selection against recombinants retains large unrecombined genomic blocks that act as strong barriers to gene flow. In the hybrid zone between the fire-bellied toads Bombina Bombina and B. variegata (Anura: Bombinatoridae), two anciently diverged and ecologically distinct taxa meet and produce abundant, fertile hybrids. The dense linkage map presented here enables genomic analysis of the selection-recombination balance that keeps the two gene pools from merging into one. We mapped 4,775 newly developed marker loci from bait-enriched genomic libraries in F2 crosses. The enrichment targets were selected from a draft assembly of the B. variegata genome, after filtering highly repetitive sequences. We developed a novel approach to infer the most likely diplotype per sample and locus from the raw read mapping data, which is robust to over-merging and obviates arbitrary filtering thresholds. Large-scale synteny between Bombina and Xenopus tropicalis supports the resulting linkage map. By assessing the sex of late-stage F2 tadpoles from histological sections, we also identified the sex-determining region in the Bombina genome to 7 cM on LG5, which is homologous to X. tropicalis chromosome 5, and inferred male heterogamety, suggestive of an XY sex determination mechanism. Interestingly, chromosome 5 has been repeatedly recruited as a sex chromosome in anurans with XY sex determination.
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The structure of spermatogenic cysts and number of Sertoli cells in the testes of Bombina Bombina and Bombina variegata (Bombinatoridae, Anura, Amphibia)
Zoomorphology, 2017Co-Authors: Beata Rozenblut-kościsty, Jacek M Szymura, Anna Pecio, Rafał Piprek, Jolanta Bartmańska, Maria OgielskaAbstract:Spermatogenesis in amphibians occurs inside spermatogenic cysts formed by Sertoli cells (SCs). SCs do not constitute a permanent population and their number increases during spermatogenic cycle and ontogeny. The number of SCs per cyst is strongly correlated with the number of germ cells in the same cyst and with the cyst volume. The number of germ and Sertoli cells and cyst volumes are species-specific and increase along with age. The mean number of primary spermatocytes per one Sertoli cell (efficiency) was the same, irrespective of age and species. Late spermatids are not attached to the Sertoli cells and do not form bundles as in other amphibians. The number of primary spermatocytes inside a cyst was the yield of several mitotic cycles of a single primary spermatogonium (stem cell) and its descendants, i.e., secondary spermatogonia. In adult Bombina Bombina , the majority of cysts contained germ cells after 6–8 cycles, in juveniles after 4–6 cycles, and in adult Bombina variegata after 5–7 cycles. The mean numbers of SCs per cyst were 27.4 ± 9.9 in adult B. Bombina , 6.42 ± 2.32 in adult B. variegata , and 7.1 ± 2.33 in juvenile B. Bombina . The number of primary spermatocytes per one SC (SC efficiency) in B. Bombina in adults was 6.5 ± 2.2, in juveniles 9.0 ± 4.2, and in adult B. variegata 7.40 ± 2.96. Spermatogenesis in Bombinatoridae is exceptional owing to the lack of intimate contact between SCs and germ cells.
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para allopatry in hybridizing fire bellied toads Bombina Bombina and b variegata inference from transcriptome wide coalescence analyses
Evolution, 2016Co-Authors: Beate Nurnberger, Konrad Lohse, Anna Fijarczyk, Jacek M Szymura, Mark BlaxterAbstract:Ancient origins, profound ecological divergence, and extensive hybridization make the fire-bellied toads Bombina Bombina and B. variegata (Anura: Bombinatoridae) an intriguing test case of ecological speciation. Previous modeling has proposed that the narrow Bombina hybrid zones represent strong barriers to neutral introgression. We test this prediction by inferring the rate of gene exchange between pure populations on either side of the intensively studied Krakow transect. We developed a method to extract high confidence sets of orthologous genes from de novo transcriptome assemblies, fitted a range of divergence models to these data and assessed their relative support with analytic likelihood calculations. There was clear evidence for postdivergence gene flow, but, as expected, no perceptible signal of recent introgression via the nearby hybrid zone. The analysis of two additional Bombina taxa (B. v. scabra and B. orientalis) validated our parameter estimates against a larger set of prior expectations. Despite substantial cumulative introgression over millions of years, adaptive divergence of the hybridizing taxa is essentially unaffected by their lack of reproductive isolation. Extended distribution ranges also buffer them against small-scale environmental perturbations that have been shown to reverse the speciation process in other, more recent ecotypes.
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OrthoMCL_orfs_Bor
2016Co-Authors: Beate Nurnberger, Konrad Lohse, Anna Fijarczyk, Jacek M Szymura, Mark L. BlaxterAbstract:Predicted peptide sequences from the Bombina orientalis reference transcriptom
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reference_contigs_Bor
2016Co-Authors: Beate Nurnberger, Konrad Lohse, Anna Fijarczyk, Jacek M Szymura, Mark L. BlaxterAbstract:List of contig names in the Bombina orientalis reference transcriptom
Beate Nurnberger - One of the best experts on this subject based on the ideXlab platform.
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Tadpoles of hybridising fire-bellied toads (B. Bombina and B. variegata) differ in their susceptibility to predation.
PloS one, 2020Co-Authors: Radovan Smolinský, Vojtech Baláž, Beate NurnbergerAbstract:The role of adaptive divergence in the formation of new species has been the subject of much recent debate. The most direct evidence comes from traits that can be shown to have diverged under natural selection and that now contribute to reproductive isolation. Here, we investigate differential adaptation of two fire-bellied toads (Anura, Bombinatoridae) to two types of aquatic habitat. Bombina Bombina and B. variegata are two anciently diverged taxa that now reproduce in predator-rich ponds and ephemeral aquatic sites, respectively. Nevertheless, they hybridise extensively wherever their distribution ranges adjoin. We show in laboratory experiments that, as expected, B. variegata tadpoles are at relatively greater risk of predation from dragonfly larvae, even when they display a predator-induced phenotype. These tadpoles spent relatively more time swimming and so prompted more attacks from the visually hunting predators. We argue in the discussion that genomic regions linked to high activity in B. variegata should be barred from introgression into the B. Bombina gene pool and thus contribute to gene flow barriers that keep the two taxa from merging into one.
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a dense linkage map for a large repetitive genome discovery of the sex determining region in hybridising fire bellied toads Bombina Bombina and b variegata
bioRxiv, 2020Co-Authors: Beate Nurnberger, Mark Blaxter, S J E Baird, D Cizkova, A Bryjova, Austin B Mudd, Jacek M SzymuraAbstract:Abstract Hybrid zones that result from secondary contact between diverged populations offer unparalleled insight into the genetic architecture of emerging reproductive barriers and so shed light on the process of speciation. Natural selection and recombination jointly determine their dynamics, leading to a range of outcomes from finely fragmented mixtures of the parental genomes that facilitate introgression to a situation where strong selection against recombinants retains large unrecombined genomic blocks that act as strong barriers to gene flow. In the hybrid zone between the fire-bellied toads Bombina Bombina and B. variegata (Anura: Bombinatoridae), two anciently diverged and ecologically distinct taxa meet and produce abundant, fertile hybrids. The dense linkage map presented here enables genomic analysis of the selection-recombination balance that keeps the two gene pools from merging into one. We mapped 4,775 newly developed marker loci from bait-enriched genomic libraries in F2 crosses. The enrichment targets were selected from a draft assembly of the B. variegata genome, after filtering highly repetitive sequences. We developed a novel approach to infer the most likely diplotype per sample and locus from the raw read mapping data, which is robust to over-merging and obviates arbitrary filtering thresholds. Large-scale synteny between Bombina and Xenopus tropicalis supports the resulting linkage map. By assessing the sex of late-stage F2 tadpoles from histological sections, we also identified the sex-determining region in the Bombina genome to 7 cM on LG5, which is homologous to X. tropicalis chromosome 5, and inferred male heterogamety, suggestive of an XY sex determination mechanism. Interestingly, chromosome 5 has been repeatedly recruited as a sex chromosome in anurans with XY sex determination.
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para allopatry in hybridizing fire bellied toads Bombina Bombina and b variegata inference from transcriptome wide coalescence analyses
Evolution, 2016Co-Authors: Beate Nurnberger, Konrad Lohse, Anna Fijarczyk, Jacek M Szymura, Mark BlaxterAbstract:Ancient origins, profound ecological divergence, and extensive hybridization make the fire-bellied toads Bombina Bombina and B. variegata (Anura: Bombinatoridae) an intriguing test case of ecological speciation. Previous modeling has proposed that the narrow Bombina hybrid zones represent strong barriers to neutral introgression. We test this prediction by inferring the rate of gene exchange between pure populations on either side of the intensively studied Krakow transect. We developed a method to extract high confidence sets of orthologous genes from de novo transcriptome assemblies, fitted a range of divergence models to these data and assessed their relative support with analytic likelihood calculations. There was clear evidence for postdivergence gene flow, but, as expected, no perceptible signal of recent introgression via the nearby hybrid zone. The analysis of two additional Bombina taxa (B. v. scabra and B. orientalis) validated our parameter estimates against a larger set of prior expectations. Despite substantial cumulative introgression over millions of years, adaptive divergence of the hybridizing taxa is essentially unaffected by their lack of reproductive isolation. Extended distribution ranges also buffer them against small-scale environmental perturbations that have been shown to reverse the speciation process in other, more recent ecotypes.
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OrthoMCL_orfs_Bor
2016Co-Authors: Beate Nurnberger, Konrad Lohse, Anna Fijarczyk, Jacek M Szymura, Mark L. BlaxterAbstract:Predicted peptide sequences from the Bombina orientalis reference transcriptom
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reference_contigs_Bor
2016Co-Authors: Beate Nurnberger, Konrad Lohse, Anna Fijarczyk, Jacek M Szymura, Mark L. BlaxterAbstract:List of contig names in the Bombina orientalis reference transcriptom
Ralph Tiedemann - One of the best experts on this subject based on the ideXlab platform.
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Introgression of Austrian fire-bellied toads (Bombina Bombina) into northern German populations confirmed by complete mitochondrial genomes and transcriptome-wide Single Nucleotide Polymorphisms (SNPs)
2019Co-Authors: Binia De Cahsan, Michael V. Westbury, Hauke Drews, Günter Gollmann, Sofia Paraskevopoulou, Moritz Ott, Ralph TiedemannAbstract:Abstract The fire-bellied toad (Bombina Bombina) is a small amphibian species inhabiting the lowlands of Central and Eastern Europe. Continual population fragmentation has caused a decrease in genetic diversity and gene flow in margin populations in Northern Germany. Previous research using mitochondrial control region data uncovered a translocation of allochthonous toads from Austria into a single northern German population, Hogsdorf. Microsatellite and MHC data also confirmed this introgression, but were limited in revealing the true extent of this translocation for all investigated populations. Here, we utilize complete mitochondrial genomes and nuclear data, in the form of transcriptomes, to unravel the full extent of admixture from this translocation. The increased number of loci enabled us to uncover signs of introgression in four out of the five northern populations investigated. Moreover, as hybrids between B. Bombina and its sister species, B. variegata, have been reported in Austria, we also investigated whether B. variegata alleles were translocated into the northern gene pool. We found evidence for hybridization between Austrian B. Bombina and B. variegata, as well as traces of B. variegata alleles in one of the five German populations included in this study. These findings present the first reported case of introgressed relict B. variegata alleles from the Southern lineage being detected in Northern Germany.
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The complete mitochondrial genome of a European fire-bellied toad (Bombina Bombina) from Germany
Mitochondrial DNA Part B, 2019Co-Authors: Binia De Cahsan, Michael V. Westbury, Hauke Drews, Ralph TiedemannAbstract:AbstractThe European fire-bellied toad, Bombina Bombina, is a small aquatic toad belonging to the family Bombinatoridae. The species is native to the lowlands of Central and Eastern Europe, where p...
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Environmental niche factor analysis (ENFA) relates environmental parameters to abundance and genetic diversity in an endangered amphibian, the fire-bellied-toad (Bombina Bombina)
Conservation Genetics, 2014Co-Authors: N. Dolgener, L. Freudenberger, N. Schneeweiss, P. L. Ibisch, M. Schluck, Ralph TiedemannAbstract:Increasing attempts are made to understand the factors responsible for both the demographic and genetic depletion in amphibian populations. Landscape genetics aims at a spatially explicit correlation of genetic population parameters to landscape features. Using data from the endangered fire-bellied toad Bombina Bombina in Brandenburg (Northeastern Germany), we performed an environmental niche factor analysis (ENFA), relating demographic (abundance) and genetic (diversity at 17 microsatellite loci and partial sequences of the mitochondrial control region in 434 individuals from 16 populations) parameters to ecological and anthropogenic variables such as temperature, precipitation, soil wetness, water runoff, vegetation density, and road/traffic impact. We found significant correlations between road disturbance and observed heterozygosity and between soil wetness and mitochondrial diversity. As the influences of the environmental variables can differ between different indicators for genetic diversity, population size and abundance data, our ENFA-based landscape genetics approach allows us to put various aspects of long- versus short term effective population size and genetic connectivity into an ecological and spatially explicit context, enabling potentially even forecast assessment under future environmental scenarios.
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Allochthonous individuals in managed populations of the fire-bellied toad Bombina Bombina: Genetic detection and conservation implications
Limnologica, 2012Co-Authors: Christiane Schröder, Hauke Drews, N. Dolgener, Ina Pokorny, Christoph Herden, Ralph TiedemannAbstract:The ongoing global amphibian decline calls for an increase of habitat and population management efforts. Pond restoration and construction is more and more accompanied by breeding and translocation programs. However, the appropriateness of translocations as a tool for conservation has been widely debated, as it can cause biodiversity loss through genetic homogenization and can disrupt local adaptation, eventually leading to outbreeding depression. In this study, we investigated the genetic structure of two translocated populations of the critically endangered fire-bellied toad Bombina Bombina at its north western distribution edge using supposedly neutral genetic markers (variation in the mitochondrial control region and microsatellites) as well as a marker under selection (major histocompatibility complex (MHC) genes). While one of the newly established populations showed the typical genetic composition of surrounding populations, the other was extremely diverged without clear affinity to its putative source. In this population we detected a profound impact of allochthonous individuals: 100% of the analyzed individuals exhibited a highly divergent mitochondrial haplotype which was otherwise found in Austria. 83% of them were also assigned to Austria by the analysis of microsatellites. Interestingly, for the adaptive marker (MHC) local alleles were predominant in this population, while only very few alleles were shared with the Austrian population. Probably Mendelian inheritance has reshuffled genotypes such that adaptive local alleles are maintained (here, MHC), while presumably neutral allochthonous alleles dominate at other loci. The release of allochthonous individuals generally increased the genetic variability of the affected population without wiping out locally adaptive genotypes. Thus, outbreeding depression might be less apparent than sometimes thought and natural selection appears strong enough to maintain locally adaptive alleles, at least in functionally important immune system genes.
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nine new tetranucleotide microsatellite markers for the fire bellied toad Bombina Bombina
Molecular Ecology Notes, 2006Co-Authors: J S Hauswaldt, Christiane Schröder, Ralph TiedemannAbstract:We describe nine new polymorphic tetranucleotide microsatellite loci isolated from the fire-bellied toad (Bombina Bombina). The relative yield of new loci was higher than described in previous studies in amphibians: out of 12 loci initially evaluated, nine were polymorphic and amplifying reliably. Number of alleles ranged from four to 10 and observed heterozygosities from 0.47 to 0.91. Seven loci were polymorphic also in Bombina variegata and five in Bombina orientalis. Enrichment protocols yielding long flanking regions potentially overcome difficulties (i.e, low yield of reliable loci relative to number of clones screened) which have been reported in microsatellite development in anurans.
Nicholas H Barton - One of the best experts on this subject based on the ideXlab platform.
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hybridization of Bombina Bombina and b variegata anura discoglossidae at a sharp ecotone in western ukraine comparisons across transects and over time
Evolution, 2006Co-Authors: Alexey Yanchukov, Jacek M Szymura, Sebastian Hofman, S V Mezhzherin, Sviatoslav Y Morozovleonov, Nicholas H Barton, Beate NurnbergerAbstract:Bombina Bombina and B. variegata are two anciently diverged toad taxa that have adapted to different breeding habitats yet hybridize freely in zones of overlap where their parapatric distributions meet. Here, we report on a joint genetic and ecological analysis of a hybrid zone in the vicinity of Stryi in western Ukraine. We used five unlinked allozyme loci, two nuclear single nucleotide polymorphisms and a mitochondrial DNA haplotype as genetic markers. Parallel allele frequency clines with a sharp central step occur across a sharp ecotone, where transitions in aquatic habitat, elevation, and terrestrial vegetation coincide. The width of the hybrid zone, estimated as the inverse of the maximum gradient in allele frequency, is 2.3 km. This is the smallest of four estimates derived from different clinal transects across Europe. We argue that the narrow cline near Stryi is mainly due to a combination of habitat distribution and habitat preference. Adult toads show a preference for either ponds (B. Bombina) or puddles (B. variegata), which is known to affect the distribution of genotypes within the hybrid zones. At Stryi, it should cause a reduction of the dispersal rate across the ecotone and thus narrow the cline. A detailed comparison of all five intensively studied Bombina transects lends support to the hypothesis that habitat distribution plus habitat preference can jointly affect the structure of hybrid zones and, ultimately, the resulting barriers to gene flow between differentiated gene pools. This study also represents a resampling of an area that was last studied more than 70 years ago. Our allele-frequency clines largely coincide with those that were described then on the basis of morphological variation. However, we found asymmetrical introgression of B. variegata genes into B. Bombina territory along the bank of a river.
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the maintenance of reproductive isolation in a mosaic hybrid zone between the fire bellied toads Bombina Bombina and b variegata
Evolution, 2003Co-Authors: Nicholas H Barton, Catriona J Maccallum, Timothy H Vines, S C Kohler, M Thiel, Ioan Ghira, T R Sands, Beate NurnbergerAbstract:Mosaic hybrid zones arise when ecologically differentiated taxa hybridize across a network of habitat patches. Frequent interbreeding across a small-scale patchwork can erode species differences that might have been preserved in a clinal hybrid zone. In particular, the rapid breakdown of neutral divergence sets an upper limit to the time for which differences at marker loci can persist. We present here a case study of a mosaic hybrid zone between the fire-bellied toads Bombina Bombina and B. variegata (Anura: Discoglossidae) near Apahida in Romania. In our 20 3 20 km study area, we detected no evidence of a clinal transition but found a strong association between aquatic habitat and mean allele frequencies at four molecular markers. In particular, pure populations of B. Bombina in ponds appear to cause massive introgression into the surrounding B. variegata gene pool found in temporary aquatic sites. Nevertheless, the genetic structure of these hybrid populations was remarkably similar to those of a previously studied transect near Pescenica (Croatia), which had both clinal and mosaic features: estimates of heterozygote deficit and linkage disequilibrium in each country are similar. In Apahida, the observed strong linkage disequilibria should stem from an imperfect habitat preference that guides most (but not all) adults into the habitats to which they are adapted. In the absence of a clinal structure, the inferred migration rate between habitats implies that associations between selected loci and neutral markers should break down rapidly. Although plausible selection strengths can maintain differentiation at those loci adapting the toads to either permanent or temporary breeding sites, the divergence at neutral markers must be transient. The hybrid zone may be approaching a state in which the gene pools are homogenized at all but the selected loci, not dissimilar from an early stage of sympatric divergence.
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hybrid dysfunction in fire bellied toads Bombina
Evolution, 1999Co-Authors: Loeske E. B. Kruuk, Jason S. Gilchrist, Nicholas H BartonAbstract:Reproductive isolation between two taxa may be due to endogenous selection, which is generated by incompatibilities between the respective genomes, to exogenous selection, which is generated by differential adaptations to alternative environments, or to both. The continuing debate over the relative importance of either mode of selection has highlighted the need for unambiguous data on the fitness of hybrid genotypes. The hybrid zone between the fire-bellied toad (Bombina Bombina) and the yellow-bellied toad (B. variegata) in central Europe involves adaptation to different environments, but evidence of hybrid dysfunction is equivocal. In this study, we followed the development under laboratory conditions of naturally laid eggs collected from a transect across the Bombina hybrid zone in Croatia. Fitness was significantly reduced in hybrid populations: Egg batches from the center of the hybrid zone showed significantly higher embryonic and larval mortality and higher frequencies of morphological abnormalities relative to either parental type. Overall mortality from day of egg collection to three weeks after hatching reached 20% in central hybrid populations, compared to 2% in pure populations. There was no significant difference in fitness between two parental types. Within hybrid populations, there was considerable variation in fitness, with some genotypes showing no evidence of reduced viability. We discuss the implications of these findings for our understanding of barriers to gene flow between species.
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habitat preference in the Bombina hybrid zone in croatia
Evolution, 1998Co-Authors: Catriona J Maccallum, Beate Nurnberger, Nicholas H Barton, Jacek M SzymuraAbstract:This paper demonstrates the effect of habitat heterogeneity and a habitat preference on the genetic structure of a hybrid zone between the toads Bombina Bombina and B. variegata (Anura: Discoglossidae); 1613 toads from 85 sites across a transect near Pescenica, Croatia, were scored for five unlinked diagnostic allozyme markers. These were found to be largely concordant. Aside from minor systematic deviations, there was little variance in allele frequency among loci within sites. Yet the allele frequencies did not follow a smooth cline, but formed a mosaic in the center, such that neighboring sites could differ markedly in their enzyme score. A detailed ecological survey revealed a correlation between this pattern and habitat. In keeping with the typical breeding sites of the parental taxa, B. Bombina-like hybrids were found more often in ponds, whereas B. variegata-like hybrids were more common in puddles. In addition, there was significant heterozygote deficit (FIS) and strong linkage disequilibrium (R), both of which were stronger on the B. Bombina side of the transect, and stronger in puddles than ponds. Mark-recapture data showed: (1) that the animals disperse beyond the scale of the habitat pattern; (2) frequent turn-over of individuals within sites; and (3) nonrandom movement between two sites of different habitat type. We conclude that an active habitat preference must contribute to the observed association between marker alleles and habitat. As a consequence, there is incomplete mixing of the two gene pools, which could explain the high level of FIS and R. The asymmetry in these parameters may reflect asymmetry in the preference or in the distribution of habitats across the zone. We discuss the implications of habitat preference for the dynamics of hybrid zones.
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natural selection on quantitative traits in the Bombina hybrid zone
Evolution, 1995Co-Authors: Beate Nurnberger, Jason S. Gilchrist, Nicholas H Barton, Catriona J Maccallum, M C ApplebyAbstract:Observations on the means, variances, and covariances of quantitative traits across hybrid zones can give information similar to that from Mendelian markers. In addition, they can identify particular traits through which the cline is maintained. We describe a survey of six traits across the hybrid zone between Bombina Bombina and Bombina variegata (Amphibia: Discoglossidae) near Pescenica in Croatia. We obtained laboratory measuments of the belly pattern, skin thickness, mating call, skeletal form, egg size, and the developmental time of tadpoles. Although offspring from hybrid populations showed no evidence of reduced viability, a third of the F1 families failed completely, irrespective of the direction of the cross. All traits differed significantly between the taxa. Clines in belly pattern, skin thickness, mating call, and skeletal form were closely concordant with clines in four diagnostic enzyme loci. However, the cline in developmental time was displaced towards Bombina, and the cline in egg size was displaced towards variegata. This discordance could be because the traits are not inherited additively or because they are subject to different selection pressures. We favor the latter explanation in the case of developmental time. We show that moderate selection acting directly on a trait suffices to shift its position; rather stronger selection is needed to change its width appreciably. Within hybrid populations, there are significant associations among quantitative traits, and between traits and enzymes. Phenotypic variances also increase in hybrid populations. These observations can be explained by linkage disequilibria among the underlying loci. However, the average magnitude of the covariance between traits is about half that expected from the linkage disequilibria between enzyme loci. The discrepancy is not readily explained by nonadditive gene action. This puzzle is now unresolved and calls for further investigation.
Tianbao Chen - One of the best experts on this subject based on the ideXlab platform.
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Identification and molecular cloning of novel trypsin inhibitor analogs from the dermal venom of the Oriental fire-bellied toad (Bombina orientalis) and the European yellow-bellied toad (Bombina variegata).
Peptides, 2003Co-Authors: Tianbao Chen, Christopher ShawAbstract:The structural diversity of polypeptides in amphibian skin secretion probably reflects different roles in dermal regulation or in defense against predators. Here we report the structures of two novel trypsin inhibitor analogs, BOTI and BVTI, from the dermal venom of the toads, Bombina orientalis and Bombina variegata. Cloning of their respective precursors was achieved from lyophilized venom cDNA libraries for the first time. Amino acid alignment revealed that both deduced peptides, consisting of 60 amino acid residues, including 10 cysteines and the reactive center motif, -CDKKC-, can be affirmed as structural homologs of the trypsin inhibitor from Bombina Bombina skin.
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novel bradykinins and their precursor cdnas from european yellow bellied toad Bombina variegata skin
FEBS Journal, 2002Co-Authors: Tianbao Chen, David F Orr, Anthony J Bjourson, Stephen Mcclean, Martin Orourke, David Hirst, Pingfan Rao, C ShawAbstract:Two novel bradykinin-related peptides (Ala3,Thr6)-bradykinin and (Val1,Thr3,Thr6)-bradykinin, were identified by a systematic sequencing study of peptides in the defensive skin secretion of the yellow-bellied toad, Bombina variegata. These peptides are the first amphibian skin bradykinins to exhibit amino acid substitutions at the Pro3 position of the bradykinin nonapeptide. Previously reported bradykinins from other Bombina species were not detected. Respective precursor cDNAs, designated BVK-1 and BVK-2, respectively, were cloned from a skin library by 3'- and 5'-RACE reactions. BVK-1 contained an open-reading frame of 97 amino acids encoding a single copy of (Ala3,Thr6)-bradykinin and similarly, the open-reading frame of BVK-2 consisted of 96 amino acids encoding a single copy of (Val1,Thr3,Thr6)-bradykinin. Synthetic replicates of each novel bradykinin were found to be active on mammalian arterial and small intestinal smooth muscle preparations. The structural diversity of bradykinins in amphibian defensive skin secretions may be related to defence against specific predators.