Federico G. Hoffmann

Comparative genomics reveals insights into avian genome evolution and adaptation

Birds are the most species-rich class of tetrapod vertebrates and have wide relevance across many research fields. We explored bird macroevolution using full genomes from 48 avian species representing all major extant clades. The avian genome is principally characterized by its constrained size, which predominantly arose because of lineage-specific erosion of repetitive elements, large segmental deletions, and gene loss. Avian genomes furthermore show a remarkably high degree of evolutionary stasis at the levels of nucleotide sequence, gene synteny, and chromosomal structure. Despite this pattern of conservation, we detected many non-neutral evolutionary changes in protein-coding genes and noncoding regions. These analyses reveal that pan-avian genomic diversity covaries with adaptations to different lifestyles and convergent evolution of traits.

The Burmese python genome reveals the molecular basis for extreme adaptation in snakes

Significance The molecular basis of morphological and physiological adaptations in snakes is largely unknown. Here, we study these phenotypes using the genome of the Burmese python (Python molurus bivittatus), a model for extreme phenotypic plasticity and metabolic adaptation. We discovered massive rapid changes in gene expression that coordinate major changes in organ size and function after feeding. Many significantly responsive genes are associated with metabolism, development, and mammalian diseases. A striking number of genes experienced positive selection in ancestral snakes. Such genes were related to metabolism, development, lungs, eyes, heart, kidney, and skeletal structure—all highly modified features in snakes. Snake phenotypic novelty seems to be driven by the system-wide coordination of protein adaptation, gene expression, and changes in genome structure. Snakes possess many extreme morphological and physiological adaptations. Identification of the molecular basis of these traits can provide novel understanding for vertebrate biology and medicine. Here, we study snake biology using the genome sequence of the Burmese python (Python molurus bivittatus), a model of extreme physiological and metabolic adaptation. We compare the python and king cobra genomes along with genomic samples from other snakes and perform transcriptome analysis to gain insights into the extreme phenotypes of the python. We discovered rapid and massive transcriptional responses in multiple organ systems that occur on feeding and coordinate major changes in organ size and function. Intriguingly, the homologs of these genes in humans are associated with metabolism, development, and pathology. We also found that many snake metabolic genes have undergone positive selection, which together with the rapid evolution of mitochondrial proteins, provides evidence for extensive adaptive redesign of snake metabolic pathways. Additional evidence for molecular adaptation and gene family expansions and contractions is associated with major physiological and phenotypic adaptations in snakes; genes involved are related to cell cycle, development, lungs, eyes, heart, intestine, and skeletal structure, including GRB2-associated binding protein 1, SSH, WNT16, and bone morphogenetic protein 7. Finally, changes in repetitive DNA content, guanine-cytosine isochore structure, and nucleotide substitution rates indicate major shifts in the structure and evolution of snake genomes compared with other amniotes. Phenotypic and physiological novelty in snakes seems to be driven by system-wide coordination of protein adaptation, gene expression, and changes in the structure of the genome.

Three crocodilian genomes reveal ancestral patterns of evolution among archosaurs

INTRODUCTION Crocodilians and birds are the two extant clades of archosaurs, a group that includes the extinct dinosaurs and pterosaurs. Fossils suggest that living crocodilians (alligators, crocodiles, and gharials) have a most recent common ancestor 80 to 100 million years ago. Extant crocodilians are notable for their distinct morphology, limited intraspecific variation, and slow karyotype evolution. Despite their unique biology and phylogenetic position, little is known about genome evolution within crocodilians. Evolutionary rates of tetrapods inferred from DNA sequences anchored by ultraconserved elements. Evolutionary rates among reptiles vary, with especially low rates among extant crocodilians but high rates among squamates. We have reconstructed the genomes of the common ancestor of birds and of all archosaurs (shown in gray silhouette, although the morphology of these species is uncertain). RATIONALE Genome sequences for the American alligator, saltwater crocodile, and Indian gharial—representatives of all three extant crocodilian families—were obtained to facilitate better understanding of the unique biology of this group and provide a context for studying avian genome evolution. Sequence data from these three crocodilians and birds also allow reconstruction of the ancestral archosaurian genome. RESULTS We sequenced shotgun genomic libraries from each species and used a variety of assembly strategies to obtain draft genomes for these three crocodilians. The assembled scaffold N50 was highest for the alligator (508 kilobases). Using a panel of reptile genome sequences, we generated phylogenies that confirm the sister relationship between crocodiles and gharials, the relationship with birds as members of extant Archosauria, and the outgroup status of turtles relative to birds and crocodilians. We also estimated evolutionary rates along branches of the tetrapod phylogeny using two approaches: ultraconserved element–anchored sequences and fourfold degenerate sites within stringently filtered orthologous gene alignments. Both analyses indicate that the rates of base substitution along the crocodilian and turtle lineages are extremely low. Supporting observations were made for transposable element content and for gene family evolution. Analysis of whole-genome alignments across a panel of reptiles and mammals showed that the rate of accumulation of micro-insertions and microdeletions is proportionally lower in crocodilians, consistent with a single underlying cause of a reduced rate of evolutionary change rather than intrinsic differences in base repair machinery. We hypothesize that this single cause may be a consistently longer generation time over the evolutionary history of Crocodylia. Low heterozygosity was observed in each genome, consistent with previous analyses, including the Chinese alligator. Pairwise sequential Markov chain analysis of regional heterozygosity indicates that during glacial cycles of the Pleistocene, each species suffered reductions in effective population size. The reduction was especially strong for the American alligator, whose current range extends farthest into regions of temperate climates. CONCLUSION We used crocodilian, avian, and outgroup genomes to reconstruct 584 megabases of the archosaurian common ancestor genome and the genomes of key ancestral nodes. The estimated accuracy of the archosaurian genome reconstruction is 91% and is higher for conserved regions such as genes. The reconstructed genome can be improved by adding more crocodilian and avian genome assemblies and may provide a unique window to the genomes of extinct organisms such as dinosaurs and pterosaurs. To provide context for the diversification of archosaurs—the group that includes crocodilians, dinosaurs, and birds—we generated draft genomes of three crocodilians: Alligator mississippiensis (the American alligator), Crocodylus porosus (the saltwater crocodile), and Gavialis gangeticus (the Indian gharial). We observed an exceptionally slow rate of genome evolution within crocodilians at all levels, including nucleotide substitutions, indels, transposable element content and movement, gene family evolution, and chromosomal synteny. When placed within the context of related taxa including birds and turtles, this suggests that the common ancestor of all of these taxa also exhibited slow genome evolution and that the comparatively rapid evolution is derived in birds. The data also provided the opportunity to analyze heterozygosity in crocodilians, which indicates a likely reduction in population size for all three taxa through the Pleistocene. Finally, these data combined with newly published bird genomes allowed us to reconstruct the partial genome of the common ancestor of archosaurs, thereby providing a tool to investigate the genetic starting material of crocodilians, birds, and dinosaurs.

Sequencing three crocodilian genomes to illuminate the evolution of archosaurs and amniotes

The International Crocodilian Genomes Working Group (ICGWG) will sequence and assemble the American alligator (Alligator mississippiensis), saltwater crocodile (Crocodylus porosus) and Indian gharial (Gavialis gangeticus) genomes. The status of these projects and our planned analyses are described.

Comparative phylogeography of short-tailed bats ( Carollia : Phyllostomidae)

This is the first study of comparative phylogeography involving closely related species of Neotropical bats of the family Phyllostomidae. We compared patterns of geographical variation within the five species of fruit‐eating bats currently recognized in the genus Carollia using the complete mitochondrial cytochrome‐b gene. Our results suggest that the combined effect of the uplift of the Andes and the Panamanian land bridge has been as important for bats as for terrestrial mammals in shaping present‐day biodiversity in the New World tropics. Species in this genus can be arranged in two highly supported clades, with a deep subdivision within each that corresponds well to differences across the Andes. We found three congruent phylogeographical patterns across species in this genus. First, the closer relationship between samples from western Ecuador and those from Central America, compared with populations east of the Andes in C. brevicauda, C. castanea and C. perspicillata. Second, the likelihood of a similar timing in South America for the arrival and diversification of C. brevicauda and C. perspicillata from their Central America ancestors. Third, the expansion of C. perspicillata and C. sowelli into northwestern Central America in the relatively recent past. Using a molecular clock, with rates ranging from 2.3 to 5% per 106 years, diversification within Carollia would have occurred over the last 1–4.5 Myr. These estimates agree well with the last rise of the Northern Andes and the Panama isthmus.

Whole-genome duplications spurred the functional diversification of the globin gene superfamily in vertebrates

It has been hypothesized that two successive rounds of whole-genome duplication (WGD) in the stem lineage of vertebrates provided genetic raw materials for the evolutionary innovation of many vertebrate-specific features. However, it has seldom been possible to trace such innovations to specific functional differences between paralogous gene products that derive from a WGD event. Here, we report genomic evidence for a direct link between WGD and key physiological innovations in the vertebrate oxygen transport system. Specifically, we demonstrate that key globin proteins that evolved specialized functions in different aspects of oxidative metabolism (hemoglobin, myoglobin, and cytoglobin) represent paralogous products of two WGD events in the vertebrate common ancestor. Analysis of conserved macrosynteny between the genomes of vertebrates and amphioxus (subphylum Cephalochordata) revealed that homologous chromosomal segments defined by myoglobin + globin-E, cytoglobin, and the α-globin gene cluster each descend from the same linkage group in the reconstructed proto-karyotype of the chordate common ancestor. The physiological division of labor between the oxygen transport function of hemoglobin and the oxygen storage function of myoglobin played a pivotal role in the evolution of aerobic energy metabolism, supporting the hypothesis that WGDs helped fuel key innovations in vertebrate evolution.

Gene duplication, genome duplication, and the functional diversification of vertebrate globins.

The functional diversification of the vertebrate globin gene superfamily provides an especially vivid illustration of the role of gene duplication and whole-genome duplication in promoting evolutionary innovation. For example, key globin proteins that evolved specialized functions in various aspects of oxidative metabolism and oxygen signaling pathways (hemoglobin [Hb], myoglobin [Mb], and cytoglobin [Cygb]) trace their origins to two whole-genome duplication events in the stem lineage of vertebrates. The retention of the proto-Hb and Mb genes in the ancestor of jawed vertebrates permitted a physiological division of labor between the oxygen-carrier function of Hb and the oxygen-storage function of Mb. In the Hb gene lineage, a subsequent tandem gene duplication gave rise to the proto α- and β-globin genes, which permitted the formation of multimeric Hbs composed of unlike subunits (α(2)β(2)). The evolution of this heteromeric quaternary structure was central to the emergence of Hb as a specialized oxygen-transport protein because it provided a mechanism for cooperative oxygen-binding and allosteric regulatory control. Subsequent rounds of duplication and divergence have produced diverse repertoires of α- and β-like globin genes that are ontogenetically regulated such that functionally distinct Hb isoforms are expressed during different stages of prenatal development and postnatal life. In the ancestor of jawless fishes, the proto Mb and Hb genes appear to have been secondarily lost, and the Cygb homolog evolved a specialized respiratory function in blood-oxygen transport. Phylogenetic and comparative genomic analyses of the vertebrate globin gene superfamily have revealed numerous instances in which paralogous globins have convergently evolved similar expression patterns and/or similar functional specializations in different organismal lineages.

Gene cooption and convergent evolution of oxygen transport hemoglobins in jawed and jawless vertebrates

Natural selection often promotes evolutionary innovation by coopting preexisting genes for new functions, and this process may be greatly facilitated by gene duplication. Here we report an example of cooptive convergence where paralogous members of the globin gene superfamily independently evolved a specialized O2 transport function in the two deepest branches of the vertebrate family tree. Specifically, phylogenetic evidence demonstrates that erythroid-specific O2 transport hemoglobins evolved independently from different ancestral precursor proteins in jawed vertebrates (gnathostomes) and jawless fish (cyclostomes, represented by lamprey and hagfish). A comprehensive phylogenetic analysis of the vertebrate globin gene superfamily revealed that the erythroid hemoglobins of cyclostomes are orthologous to the cytoglobin protein of gnathostome vertebrates, a hexacoordinate globin that has no O2 transport function and that is predominantly expressed in fibroblasts and related cell types. The phylogeny reconstruction also revealed that vertebrate-specific globins are grouped into four main clades: (i) cyclostome hemoglobin + cytoglobin, (ii) myoglobin + globin E, (iii) globin Y, and (iv) the α- and β-chain hemoglobins of gnathostomes. In the hemoglobins of gnathostomes and cyclostomes, multisubunit quaternary structures provide the basis for cooperative O2 binding and allosteric regulation by coupling the effects of ligand binding at individual subunits with interactions between subunits. However, differences in numerous structural details belie their independent origins. This example of convergent evolution of protein function provides an impressive demonstration of the ability of natural selection to cobble together complex design solutions by tinkering with different variations of the same basic protein scaffold.

Differential loss of embryonic globin genes during the radiation of placental mammals

The differential gain and loss of genes from homologous gene families represents an important source of functional variation among the genomes of different species. Differences in gene content between species are primarily attributable to lineage-specific gene gains via duplication and lineage-specific losses via deletion or inactivation. Here, we use a comparative genomic approach to investigate this process of gene turnover in the β-globin gene family of placental mammals. By analyzing genomic sequence data from representatives of each of the main superordinal clades of placental mammals, we were able to reconstruct pathways of gene family evolution during the basal radiation of this physiologically and morphologically diverse vertebrate group. Our analysis revealed that an initial expansion of the nonadult portion of the β-globin gene cluster in the ancestor of placental mammals was followed by the differential loss and retention of ancestral gene lineages, thereby generating variation in the complement of embryonic globin genes among contemporary species. The sorting of ε-, γ-, and η-globin gene lineages among the basal clades of placental mammals has produced species differences in the functional types of hemoglobin isoforms that can be synthesized during the course of embryonic development.

Rapid Rates of Lineage-Specific Gene Duplication and Deletion in the α-Globin Gene Family

Phylogeny reconstructions of the globin gene families have revealed that paralogous genes within species are often more similar to one another than they are to their orthologous counterparts in closely related species. This pattern has been previously attributed to mechanisms of concerted evolution such as interparalog gene conversion that homogenize sequence variation between tandemly duplicated genes and therefore create the appearance of recent common ancestry. Here we report a comparative genomic analysis of the alpha-globin gene family in mammals that reveal a surprisingly high rate of lineage-specific gene duplication and deletion via unequal crossing-over. Results of our analysis reveal that patterns of sequence similarity between paralogous alpha-like globin genes from the same species are only partly explained by concerted evolution between preexisting gene duplicates. In a number of cases, sequence similarity between paralogous sequences from the same species is attributable to recent ancestry between the products of de novo gene duplications. As a result of this surprisingly rapid rate of gene gain and loss, many mammals possess alpha-like globin genes that have no orthologous counterparts in closely related species. The resultant variation in gene copy number among species may represent an important source of regulatory variation that affects physiologically important aspects of blood oxygen transport and aerobic energy metabolism.