RESUMO
Frogs are an ecologically diverse and phylogenetically ancient group of anuran amphibians that include important vertebrate cell and developmental model systems, notably the genus Xenopus. Here we report a high-quality reference genome sequence for the western clawed frog, Xenopus tropicalis, along with draft chromosome-scale sequences of three distantly related emerging model frog species, Eleutherodactylus coqui, Engystomops pustulosus, and Hymenochirus boettgeri. Frog chromosomes have remained remarkably stable since the Mesozoic Era, with limited Robertsonian (i.e., arm-preserving) translocations and end-to-end fusions found among the smaller chromosomes. Conservation of synteny includes conservation of centromere locations, marked by centromeric tandem repeats associated with Cenp-a binding surrounded by pericentromeric LINE/L1 elements. This work explores the structure of chromosomes across frogs, using a dense meiotic linkage map for X. tropicalis and chromatin conformation capture (Hi-C) data for all species. Abundant satellite repeats occupy the unusually long (~20 megabase) terminal regions of each chromosome that coincide with high rates of recombination. Both embryonic and differentiated cells show reproducible associations of centromeric chromatin and of telomeres, reflecting a Rabl-like configuration. Our comparative analyses reveal 13 conserved ancestral anuran chromosomes from which contemporary frog genomes were constructed.
Assuntos
Cromatina , Evolução Molecular , Animais , Cromatina/genética , Genoma/genética , Anuros/genética , Xenopus/genética , Centrômero/genéticaRESUMO
BACKGROUND: Animals with polyploid, hybrid nuclei offer a challenge for models of gene expression and regulation during embryogenesis. To understand how such organisms proceed through development, we examined the timing and prevalence of mortality among embryos of unisexual salamanders in the genus Ambystoma. RESULTS: Our regional field surveys suggested that heightened rates of embryo mortality among unisexual salamanders begin in the earliest stages of embryogenesis. Although we expected elevated mortality after zygotic genome activation in the blastula stage, this is not what we found among embryos which we reared in the laboratory. Once embryos entered the first cleavage stage, we found no difference in mortality rates between unisexual salamanders and their bisexual hosts. Our results are consistent with previous studies showing high rates of unisexual mortality, but counter to reports that heightened embryo mortality continues throughout embryo development. CONCLUSIONS: Possible causes of embryonic mortality in early embryogenesis suggested by our results include abnormal maternal loading of RNA during meiosis and barriers to insemination. The surprising survival rates of embryos post-cleavage invites further study of how genes are regulated during development in such polyploid hybrid organisms.
Assuntos
Urodelos/embriologia , Urodelos/genética , Animais , Desenvolvimento Embrionário , Poliploidia , Análise de Sobrevida , Urodelos/crescimento & desenvolvimentoRESUMO
The skin secretions of many frogs have genetically-encoded, endogenous antimicrobial peptides (AMPs). Other species, especially aposematic poison frogs, secrete exogenously derived alkaloids that serve as potent defense molecules. The origins of these defense systems are not clear, but a novel bile-acid derived metabolite, tauromantellic acid, was recently discovered and shown to be endogenous in poison frogs (Mantella, Dendrobates, and Epipedobates). These observations raise questions about the evolutionary history of AMP genetic elements, the mechanism and function of tauromatellic acid production, and links between these systems. To understand the diversity and expression of AMPs among frogs, we assembled skin transcriptomes of 13 species across the anuran phylogeny. Our analyses revealed a diversity of AMPs and AMP expression levels across the phylogenetic history of frogs, but no observations of AMPs in Mantella We examined genes expressed in the bile-acid metabolic pathway and found that CYP7A1 (Cytochrome P450), BAAT (bile acid-CoA: amino acid N-acyltransferase), and AMACR (alpha-methylacyl-CoA racemase) were highly expressed in the skin of M. betsileo and either lowly expressed or absent in other frog species. In particular, CYP7A1 catalyzes the first reaction in the cholesterol catabolic pathway and is the rate-limiting step in regulation of bile acid synthesis, suggesting unique activation of the bile acid pathway in Mantella skin. The activation of the bile acid pathway in the skin of Mantella and the lack of observed AMPs fuel new questions about the evolution of defense compounds and the ectopic expression of the bile-acid pathway.
Assuntos
Proteínas de Anfíbios/genética , Peptídeos Catiônicos Antimicrobianos/genética , Anuros/genética , Ácidos e Sais Biliares/biossíntese , Transcriptoma , Proteínas de Anfíbios/metabolismo , Animais , Peptídeos Catiônicos Antimicrobianos/metabolismo , Anuros/classificação , Anuros/metabolismo , Ácidos e Sais Biliares/genética , Filogenia , Pele/metabolismoRESUMO
Amphibians inhabiting montane riparian zones in the Neotropics are particularly vulnerable to decline, but the reasons are poorly understood. Because environmental contaminants, endocrine disruption, and pathogens often figure prominently in amphibian declines it is imperative that we understand how these factors are potentially interrelated to affect montane populations. One possibility is that increased precipitation associated with global warming promotes the deposition of contaminants in montane regions. Increased exposure to contaminants, in turn, potentially elicits chronic elevations in circulating stress hormones that could contribute to montane population declines by compromising resistance to pathogens and/or production of sex steroids regulating reproduction. Here, we test this hypothesis by examining contaminant levels, stress and sex steroid levels, and nematode abundances in male drab treefrogs, Smilisca sordida, from lowland and montane populations in Costa Rica. We found no evidence that montane populations were more likely to possess contaminants (i.e., organochlorine, organophosphate and carbamate pesticides or benzidine and chlorophenoxy herbicides) than lowland populations. We also found no evidence of elevational differences in circulating levels of the stress hormone corticosterone, estradiol or progesterone. However, montane populations possessed lower androgen levels, hosted more nematode species, and had higher nematode abundances than lowland populations. Although these results suggested that nematodes contributed to lower androgens in montane populations, we were unable to detect a significant inverse relationship between nematode abundance and androgen level. Our results suggest that montane populations of this species are not at greater risk of exposure to contaminants or chronic stress, but implicate nematodes and compromised sex steroid levels as potential threats to montane populations.
Assuntos
Anuros/parasitologia , Disruptores Endócrinos/toxicidade , Poluentes Ambientais/toxicidade , Animais , Anuros/sangue , Anuros/fisiologia , Corticosterona/sangue , Costa Rica , Disruptores Endócrinos/metabolismo , Glândulas Endócrinas/efeitos dos fármacos , Glândulas Endócrinas/fisiopatologia , Poluentes Ambientais/metabolismo , Aquecimento Global , Hormônios Esteroides Gonadais/sangue , Interações Hospedeiro-Parasita , Masculino , Nematoides/isolamento & purificação , Nematoides/patogenicidade , Dinâmica Populacional , Estresse Fisiológico , Clima Tropical/efeitos adversosRESUMO
A new study provides evidence that gene transposition from sex chromosomes to autosomes is a conserved phenomenon across mammalian species that rescues dosage-sensitive genes.
Assuntos
Deleção de Genes , Mamíferos/genética , Aberrações dos Cromossomos Sexuais , Translocação Genética , Cromossomo Y/genética , Animais , HumanosRESUMO
Frog sex chromosomes offer an ideal system for advancing our understanding of genome evolution and function because of the variety of sex determination systems in the group, the diversity of sex chromosome maturation states, the ease of experimental manipulation during early development. After briefly reviewing sex chromosome biology generally, we focus on what is known about frog sex determination, sex chromosome evolution, and recent, genomics-facilitated advances in the field. In closing we highlight gaps in our current knowledge of frog sex chromosomes, and suggest priorities for future research that can advance broad knowledge of gene dose and sex chromosome evolution.
RESUMO
Accurate gene model annotation of reference genomes is critical for making them useful. The modENCODE project has improved the D. melanogaster genome annotation by using deep and diverse high-throughput data. Since transcriptional activity that has been evolutionarily conserved is likely to have an advantageous function, we have performed large-scale interspecific comparisons to increase confidence in predicted annotations. To support comparative genomics, we filled in divergence gaps in the Drosophila phylogeny by generating draft genomes for eight new species. For comparative transcriptome analysis, we generated mRNA expression profiles on 81 samples from multiple tissues and developmental stages of 15 Drosophila species, and we performed cap analysis of gene expression in D. melanogaster and D. pseudoobscura. We also describe conservation of four distinct core promoter structures composed of combinations of elements at three positions. Overall, each type of genomic feature shows a characteristic divergence rate relative to neutral models, highlighting the value of multispecies alignment in annotating a target genome that should prove useful in the annotation of other high priority genomes, especially human and other mammalian genomes that are rich in noncoding sequences. We report that the vast majority of elements in the annotation are evolutionarily conserved, indicating that the annotation will be an important springboard for functional genetic testing by the Drosophila community.
Assuntos
Biologia Computacional/métodos , Drosophila melanogaster/genética , Perfilação da Expressão Gênica , Anotação de Sequência Molecular , Transcriptoma , Animais , Análise por Conglomerados , Drosophila melanogaster/classificação , Evolução Molecular , Éxons , Feminino , Genoma de Inseto , Humanos , Masculino , Motivos de Nucleotídeos , Filogenia , Matrizes de Pontuação de Posição Específica , Regiões Promotoras Genéticas , Edição de RNA , Sítios de Splice de RNA , Splicing de RNA , Reprodutibilidade dos Testes , Sítio de Iniciação de TranscriçãoRESUMO
Young species complexes that are widespread across ecologically disparate regions offer important insights into the process of speciation because of their relevance to how local adaptation and gene flow influence diversification. We used mitochondrial DNA and up to 28 152 genomewide single nucleotide polymorphisms from polytypic barking frogs (Craugastor augusti complex) to infer phylogenetic relationships and test for the signature of introgressive hybridization among diverging lineages. Our phylogenetic reconstructions suggest (i) a rapid Pliocene-Pleistocene radiation that produced at least nine distinct lineages and (ii) that geographic features of the arid Central Mexican Plateau contributed to two independent northward expansions. Despite clear lineage differentiation (many private alleles and high between-lineage FST scores), D-statistic tests, which differentiate introgression from ancestral polymorphism, allowed us to identify two putative instances of reticulate gene flow. Partitioned D-statistics provided evidence that these events occurred in the same direction between clades but at different points in time. After correcting for geographic distance, we found that lineages involved in hybrid gene flow interactions had higher levels of genetic variation than independently evolving lineages. These findings suggest that the nature of hybrid compatibility can be conserved overlong periods of evolutionary time and that hybridization between diverging lineages may contribute to standing levels of genetic variation.
Assuntos
Anuros/genética , Fluxo Gênico , Hibridização Genética , Animais , Evolução Biológica , Núcleo Celular/genética , DNA Mitocondrial/genética , México , Modelos Genéticos , Dados de Sequência Molecular , Filogenia , Polimorfismo de Nucleotídeo Único , Análise de Sequência de DNARESUMO
Biotic and abiotic factors have been proposed to explain patterns of reproductive character displacement, but which factor is most important to character displacement of acoustic signals is not clear. Male vocalizations of the frog Pseudacris feriarum are known to undergo reproductive character displacement in areas of sympatry with P. brimleyi and P. nigrita. Despite evidence for reinforcement as an important mechanism, local adaptation via sensory drive might explain this pattern because Pseudacris breed in different habitat types and mating signals are exposed to a variety of environments. We tested the sensory drive hypothesis by playing synthesized vocalizations representing the spectrum of variation in P. feriarum at 12 different study sites. If sensory drive has occurred, then vocalizations should transmit better in the site of origin or at ecologically similar sites. We found that variation in acoustic signals did not produce better transmission in particular sites, the effect of site was uniform, and acoustic signals often transmitted better in habitats external to their origin. Ecological variation among habitats did not explain signal degradation. Our playback experiments, ecological analyses, and comparisons of different habitat types provide no support for sensory drive as a process promoting reproductive character displacement in this system. Reinforcement is the more likely primary mechanism.
Assuntos
Adaptação Fisiológica/genética , Anuros/genética , Ecossistema , Isolamento Reprodutivo , Vocalização Animal , Animais , Anuros/fisiologia , Masculino , Som , SimpatriaRESUMO
BACKGROUND: The production of multiple transcript isoforms from one gene is a major source of transcriptome complexity. RNA-Seq experiments, in which transcripts are converted to cDNA and sequenced, allow the resolution and quantification of alternative transcript isoforms. However, methods to analyze splicing are underdeveloped and errors resulting in incorrect splicing calls occur in every experiment. RESULTS: We used RNA-Seq data to develop sequencing and aligner error models. By applying these error models to known input from simulations, we found that errors result from false alignment to minor splice motifs and antisense stands, shifted junction positions, paralog joining, and repeat induced gaps. By using a series of quantitative and qualitative filters, we eliminated diagnosed errors in the simulation, and applied this to RNA-Seq data from Drosophila melanogaster heads. We used high-confidence junction detections to specifically interrogate local splicing differences between transcripts. This method out-performed commonly used RNA-seq methods to identify known alternative splicing events in the Drosophila sex determination pathway. We describe a flexible software package to perform these tasks called Splicing Analysis Kit (Spanki), available at http://www.cbcb.umd.edu/software/spanki. CONCLUSIONS: Splice-junction centric analysis of RNA-Seq data provides advantages in specificity for detection of alternative splicing. Our software provides tools to better understand error profiles in RNA-Seq data and improve inference from this new technology. The splice-junction centric approach that this software enables will provide more accurate estimates of differentially regulated splicing than current tools.
Assuntos
Processamento Alternativo/genética , Drosophila/genética , Modelos Genéticos , Análise de Sequência de RNA/métodos , Software , Animais , Sequência de Bases , Biologia Computacional , Simulação por Computador , Feminino , Perfilação da Expressão Gênica/métodos , Masculino , Dados de Sequência Molecular , Isoformas de Proteínas/genética , RNA Mensageiro/análise , RNA Mensageiro/genéticaRESUMO
DNA sequences on X chromosomes often have a faster rate of evolution when compared to similar loci on the autosomes, and well articulated models provide reasons why the X-linked mode of inheritance may be responsible for the faster evolution of X-linked genes. We analyzed microarray and RNA-seq data collected from females and males of six Drosophila species and found that the expression levels of X-linked genes also diverge faster than autosomal gene expression, similar to the "faster-X" effect often observed in DNA sequence evolution. Faster-X evolution of gene expression was recently described in mammals, but it was limited to the evolutionary lineages shortly following the creation of the therian X chromosome. In contrast, we detect a faster-X effect along both deep lineages and those on the tips of the Drosophila phylogeny. In Drosophila males, the dosage compensation complex (DCC) binds the X chromosome, creating a unique chromatin environment that promotes the hyper-expression of X-linked genes. We find that DCC binding, chromatin environment, and breadth of expression are all predictive of the rate of gene expression evolution. In addition, estimates of the intraspecific genetic polymorphism underlying gene expression variation suggest that X-linked expression levels are not under relaxed selective constraints. We therefore hypothesize that the faster-X evolution of gene expression is the result of the adaptive fixation of beneficial mutations at X-linked loci that change expression level in cis. This adaptive faster-X evolution of gene expression is limited to genes that are narrowly expressed in a single tissue, suggesting that relaxed pleiotropic constraints permit a faster response to selection. Finally, we present a conceptional framework to explain faster-X expression evolution, and we use this framework to examine differences in the faster-X effect between Drosophila and mammals.
Assuntos
Drosophila/genética , Evolução Molecular , Expressão Gênica , Genes Ligados ao Cromossomo X , Cromossomo X , Adaptação Biológica , Animais , Mecanismo Genético de Compensação de Dose , Drosophila/metabolismo , Feminino , Heterocromatina/genética , Heterocromatina/metabolismo , Masculino , FilogeniaRESUMO
BACKGROUND: Gene dosage change is a mild perturbation that is a valuable tool for pathway reconstruction in Drosophila. While it is often assumed that reducing gene dose by half leads to two-fold less expression, there is partial autosomal dosage compensation in Drosophila, which may be mediated by feedback or buffering in expression networks. RESULTS: We profiled expression in engineered flies where gene dose was reduced from two to one. While expression of most one-dose genes was reduced, the gene-specific dose responses were heterogeneous. Expression of two-dose genes that are first-degree neighbors of one-dose genes in novel network models also changed, and the directionality of change depended on the response of one-dose genes. CONCLUSIONS: Our data indicate that expression perturbation propagates in network space. Autosomal compensation, or the lack thereof, is a gene-specific response, largely mediated by interactions with the rest of the transcriptome.
Assuntos
Mecanismo Genético de Compensação de Dose , Drosophila/genética , Redes Reguladoras de Genes , Genes de Insetos , Animais , Animais Geneticamente Modificados/genética , Cromossomos de Insetos/genética , Feminino , Dosagem de Genes , Heterogeneidade Genética , Masculino , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Transcriptoma , Cromossomo X/genéticaRESUMO
X chromosomes are preferentially transmitted through females, which may favor the accumulation of X-linked alleles/genes with female-beneficial effects. Numerous studies have shown that genes with sex-biased expression are under- or over-represented on the X chromosomes of a wide variety of organisms. The patterns, however, vary between different animal species, and the causes of these differences are unresolved. Additionally, genes with sex-biased expression tend to be narrowly expressed in a limited number of tissues, and narrowly expressed genes are also non-randomly X-linked in a taxon-specific manner. It is therefore unclear whether the unique gene content of the X chromosome is the result of selection on genes with sex-biased expression, narrowly expressed genes, or some combination of the two. To address this problem, we measured sex-biased expression in multiple Drosophila species and at different developmental time points. These data were combined with available expression measurements from Drosophila melanogaster and mouse to reconcile the inconsistencies in X-chromosome content among taxa. Our results suggest that most of the differences between Drosophila and mammals are confounded by disparate data collection/analysis approaches as well as the correlation between sex bias and expression breadth. Both the Drosophila and mouse X chromosomes harbor an excess of genes with female-biased expression after controlling for the confounding factors, suggesting that the asymmetrical transmission of the X chromosome favors the accumulation of female-beneficial mutations in X-linked genes. However, some taxon-specific patterns remain, and we provide evidence that these are in part a consequence of constraints imposed by the dosage compensation mechanism in Drosophila.
Assuntos
Mecanismo Genético de Compensação de Dose , Drosophila/genética , Genes Ligados ao Cromossomo X , Ligação Genética , Cromossomo X/genética , Animais , Mapeamento Cromossômico/métodos , Drosophila/metabolismo , Etiquetas de Sequências Expressas , Feminino , Perfilação da Expressão Gênica , Genes de Insetos , Larva/genética , Larva/metabolismo , Masculino , Camundongos , Seleção Genética , Fatores Sexuais , Testículo/citologia , Testículo/metabolismo , Cromossomo X/metabolismoRESUMO
Microarrays first made the analysis of the transcriptome possible, and have produced much important information. Today, however, researchers are increasingly turning to direct high-throughput sequencing -- RNA-Seq -- which has considerable advantages for examining transcriptome fine structure -- for example in the detection of allele-specific expression and splice junctions. In this article, we discuss the relative merits of the two techniques, the inherent biases in each, and whether all of the vast body of array work needs to be revisited using the newer technology. We conclude that microarrays remain useful and accurate tools for measuring expression levels, and RNA-Seq complements and extends microarray measurements.
Assuntos
Drosophila/genética , Perfilação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Análise de Sequência com Séries de Oligonucleotídeos/métodos , Análise de Sequência de RNA/métodos , Animais , HumanosRESUMO
Drosophila melanogaster is one of the most well studied genetic model organisms; nonetheless, its genome still contains unannotated coding and non-coding genes, transcripts, exons and RNA editing sites. Full discovery and annotation are pre-requisites for understanding how the regulation of transcription, splicing and RNA editing directs the development of this complex organism. Here we used RNA-Seq, tiling microarrays and cDNA sequencing to explore the transcriptome in 30 distinct developmental stages. We identified 111,195 new elements, including thousands of genes, coding and non-coding transcripts, exons, splicing and editing events, and inferred protein isoforms that previously eluded discovery using established experimental, prediction and conservation-based approaches. These data substantially expand the number of known transcribed elements in the Drosophila genome and provide a high-resolution view of transcriptome dynamics throughout development.
Assuntos
Drosophila melanogaster/crescimento & desenvolvimento , Drosophila melanogaster/genética , Perfilação da Expressão Gênica , Regulação da Expressão Gênica no Desenvolvimento/genética , Transcrição Gênica/genética , Processamento Alternativo/genética , Animais , Sequência de Bases , Proteínas de Drosophila/genética , Drosophila melanogaster/embriologia , Éxons/genética , Feminino , Genes de Insetos/genética , Genoma de Inseto/genética , Masculino , MicroRNAs/genética , Análise de Sequência com Séries de Oligonucleotídeos , Isoformas de Proteínas/genética , Edição de RNA/genética , RNA Mensageiro/análise , RNA Mensageiro/genética , Pequeno RNA não Traduzido/análise , Pequeno RNA não Traduzido/genética , Análise de Sequência , Caracteres SexuaisRESUMO
BACKGROUND: Drosophila females commit tremendous resources to egg production and males produce some of the longest sperm in the animal kingdom. We know little about the coordinated regulation of gene expression patterns in distant somatic tissues that support the developmental cost of gamete production. RESULTS: We determined the non-gonadal gene expression patterns of Drosophila females and males with or without a germline. Our results show that germline-dependent expression in the non-gonadal soma is extensive. Interestingly, gene expression patterns and hormone titers are consistent with a hormone axis between the gonads and non-gonadal soma. CONCLUSIONS: The germline has a long-range influence on gene expression in the Drosophila sexes. We suggest that this is the result of a germline/soma hormonal axis.
Assuntos
Drosophila melanogaster/citologia , Drosophila melanogaster/genética , Perfilação da Expressão Gênica , Células Germinativas/metabolismo , Análise de Variância , Animais , Drosophila melanogaster/metabolismo , Retroalimentação Fisiológica , Feminino , Genótipo , Gônadas , Hormônios/metabolismo , Masculino , Análise de Sequência com Séries de Oligonucleotídeos , Caracteres Sexuais , Comportamento Sexual AnimalRESUMO
Extensive departures from balanced gene dose in aneuploids are highly deleterious. However, we know very little about the relationship between gene copy number and expression in aneuploid cells. We determined copy number and transcript abundance (expression) genome-wide in Drosophila S2 cells by DNA-Seq and RNA-Seq. We found that S2 cells are aneuploid for >43 Mb of the genome, primarily in the range of one to five copies, and show a male genotype ( approximately two X chromosomes and four sets of autosomes, or 2X;4A). Both X chromosomes and autosomes showed expression dosage compensation. X chromosome expression was elevated in a fixed-fold manner regardless of actual gene dose. In engineering terms, the system "anticipates" the perturbation caused by X dose, rather than responding to an error caused by the perturbation. This feed-forward regulation resulted in precise dosage compensation only when X dose was half of the autosome dose. Insufficient compensation occurred at lower X chromosome dose and excessive expression occurred at higher doses. RNAi knockdown of the Male Specific Lethal complex abolished feed-forward regulation. Both autosome and X chromosome genes show Male Specific Lethal-independent compensation that fits a first order dose-response curve. Our data indicate that expression dosage compensation dampens the effect of altered DNA copy number genome-wide. For the X chromosome, compensation includes fixed and dose-dependent components.
Assuntos
Aneuploidia , Proteínas de Drosophila/genética , Proteínas de Drosophila/metabolismo , Drosophila/genética , Drosophila/metabolismo , Animais , Western Blotting , Linhagem Celular , Imunoprecipitação da Cromatina , Hibridização Genômica Comparativa , Mecanismo Genético de Compensação de Dose/genética , Regulação da Expressão Gênica , Masculino , Análise de Sequência com Séries de Oligonucleotídeos , Interferência de RNA , Análise de Sequência de DNA , Cromossomo X/genéticaRESUMO
Understanding the general features of speciation is an important goal in evolutionary biology, and despite significant progress, several unresolved questions remain. We analyzed an extensive comparative dataset consisting of more than 1900 crosses between 92 species of toads to infer patterns of reproductive isolation. This unique dataset provides an opportunity to examine the strength of reproductive isolation, the development and sex ratios of hybrid offspring, patterns of fertility and infertility, and polyploidization in hybrids all in the context of genetic divergence between parental species. We found that the strength of intrinsic postzygotic isolation increases with genetic divergence, but relatively high levels of divergence are necessary before reproductive isolation is complete in toads. Fertilization rates were not correlated to genetic divergence, but hatching success, the number of larvae produced, and the percentage of tadpoles reaching metamorphosis were all inversely related with genetic divergence. Hybrids between species with lower levels of divergence developed to metamorphosis, while hybrids with higher levels of divergence stopped developing in gastrula and larval stages. Sex ratios of hybrid offspring were biased towards males in 70% of crosses and biased towards females in 30% of crosses. Hybrid females from crosses between closely related species were completely fertile, while approximately half (53%) of hybrid males were sterile, with sterility predicted by genetic divergence. The degree of abnormal ploidy in hybrids was positively related to genetic divergence between parental species, but surprisingly, polyploidization had no effect on patterns of asymmetrical inviability. We discuss explanations for these patterns, including the role of Haldane's rule in toads and anurans in general, and suggest mechanisms generating patterns of reproductive isolation in anurans.