Identification and characterization of amelogenin genes in monotremes, reptiles, and amphibians

Two features make the tooth an excellent model in the study of evolutionary innovations: the relative simplicity of its structure and the fact that the major tooth-forming genes have been identified in eutherian mammals. To understand the nature of the innovation at the molecular level, it is necessary to identify the homologs of tooth-forming genes in other vertebrates. As a first step toward this goal, homologs of the eutherian amelogenin gene have been cloned and characterized in selected species of monotremes (platypus and echidna), reptiles (caiman), and amphibians (African clawed toad). Comparisons of the homologs reveal that the amelogenin gene evolves quickly in the repeat region, in which numerous insertions and deletions have obliterated any similarity among the genes, and slowly in other regions. The gene organization, the distribution of hydrophobic and hydrophilic segments in the encoded protein, and several other features have been conserved throughout the evolution of the tetrapod amelogenin gene. Clones corresponding to one locus only were found in caiman, whereas the clawed toad possesses at least two amelogenin-encoding loci.

Organization of the Hox gene cluster in the grasshopper, Schistocerca gregaria

The conserved organization of the Hox genes throughout the animal kingdom has become one of the major paradigms of evolutionary developmental biology. We have examined the organization of the Hox genes of the grasshopper, Schistocerca gregaria. We find that the grasshopper Hox cluster is over 700 kb long, and is not split into equivalents of the Antennapedia complex and the bithorax complex of Drosophila melanogaster. SgDax and probably also Sgzen, the grasshopper homologues of fushi-tarazu (ftz) and Zerknüllt (zen), respectively, are also in the cluster, showing that the non-homeotic Antp-class genes, “accessory genes,” are an ancient feature of insect Hox clusters.

A test of alternative models of diversification in tropical rainforests: Ecological gradients vs. rainforest refugia

Comparison of mitochondrial and morphological divergence in eight populations of a widespread leaf-litter skink is used to determine the relative importance of geographic isolation and natural selection in generating phenotypic diversity in the Wet Tropics Rainforest region of Australia. The populations occur in two geographically isolated regions, and within each region, in two different habitats (closed rainforest and tall open forest) that span a well characterized ecological gradient. Morphological differences among ancient geographic isolates (separated for several million years, judging by their mitochondrial DNA sequence divergence) were slight, but morphological and life history differences among habitats were large and occurred despite moderate to high levels of mitochondrial gene flow. A field experiment identified avian predation as one potential agent of natural selection. These results indicate that natural selection operating across ecological gradients can be more important than geographic isolation in similar habitats in generating phenotypic diversity. In addition, our results indicate that selection is sufficiently strong to overcome the homogenizing effects of gene flow, a necessary first step toward speciation in continuously distributed populations. Because ecological gradients may be a source of evolutionary novelty, and perhaps new species, their conservation warrants greater attention. This is particularly true in tropical regions, where most reserves do not include ecological gradients and transitional habitats.

Molecular evidence for multiple origins of woodiness and a New World biogeographic connection of the Macaronesian Island endemic Pericallis (Asteraceae: Senecioneae)

The prevalence of woody species in oceanic islands has attracted the attention of evolutionary biologists for more than a century. We used a phylogeny based on sequences of the internal-transcribed spacer region of nuclear ribosomal DNA to trace the evolution of woodiness in Pericallis (Asteraceae: Senecioneae), a genus endemic to the Macaronesian archipelagos of the Azores, Madeira, and Canaries. Our results show that woodiness in Pericallis originated independently at least twice in these islands, further weakening some previous hypotheses concerning the value of this character for tracing the continental ancestry of island endemics. The same data suggest that the origin of woodiness is correlated with ecological shifts from open to species-rich habitats and that the ancestor of Pericallis was an herbaceous species adapted to marginal habitats of the laurel forest. Our results also support Pericallis as closely related to New World genera of the tribe Senecioneae.

A natural classification of the basic helix–loop–helix class of transcription factors

A natural (evolutionary) classification is provided for 242 basic helix–loop–helix (bHLH) motif-containing proteins. Phylogenetic analyses of amino acid sequences describe the patterns of evolutionary change within the motif and delimit evolutionary lineages. These evolutionary lineages represent well known functional groups of proteins and can be further arranged into five groups based on binding to DNA at the hexanucleotide E-box, the amino acid patterns in other components of the motif, and the presence/absence of a leucine zipper. The hypothesized ancestral amino acid sequence for the bHLH transcription factor family is given together with the ancestral sequences of the subgroups. It is suggested that bHLH proteins containing a leucine zipper are not a natural, monophyletic group.

A yeast genetic system for selecting small molecule inhibitors of protein–protein interactions in nanodroplets

Cellular processes are mediated by complex networks of molecular interactions. Dissection of their role most commonly is achieved by using genetic mutations that alter, for example, protein–protein interactions. Small molecules that accomplish the same result would provide a powerful complement to the genetic approach, but it generally is believed that such molecules are rare. There are several natural products, however, that illustrate the feasibility of this approach. Split-pool synthesis now provides a simple mechanical means to prepare vast numbers of complex, even natural product-like, molecules individually attached to cell-sized polymer beads. Here, we describe a genetic system compatible with split-pool synthesis that allows the detection of cell-permeable, small molecule inhibitors of protein–protein interactions in 100- to 200-nl cell culture droplets, prepared by a recently described technique that arrays large numbers of such droplets. These “nanodroplets” contain defined media, cells, and one or more beads containing ≈100 pmol of a photoreleasable small molecule and a controlled number of cells. The engineered Saccharomyces cerevisiae cells used in this study express two interacting proteins after induction with galactose whose interaction results in cell death in the presence of 5-fluoroorotic acid (inducible reverse two-hybrid assay). Disruption of the interaction by a small molecule allows growth, and the small molecule can be introduced into the system hours before induction of the toxic interaction. We demonstrate that the interaction between the activin receptor R1 and the immunophilin protein FKBP12 can be disrupted by the small molecule FK506 at nanomolar concentrations in nanodroplets. This system should provide a general method for selecting cell-permeable ligands that can be used to study the relevance of protein–protein interactions in living cells or organisms.

DIP: The Database of Interacting Proteins: 2001 update

The Database of Interacting Proteins (DIP; http://dip.doe-mbi.ucla.edu) is a database that documents experimentally determined protein–protein interactions. Since January 2000 the number of protein–protein interactions in DIP has nearly tripled to 3472 and the number of proteins to 2659. New interactive tools have been developed to aid in the visualization, navigation and study of networks of protein interactions.

The biotic crisis and the future of evolution

The biotic crisis overtaking our planet is likely to precipitate a major extinction of species. That much is well known. Not so well known but probably more significant in the long term is that the crisis will surely disrupt and deplete certain basic processes of evolution, with consequences likely to persist for millions of years. Distinctive features of future evolution could include a homogenization of biotas, a proliferation of opportunistic species, a pest-and-weed ecology, an outburst of speciation among taxa that prosper in human-dominated ecosystems, a decline of biodisparity, an end to the speciation of large vertebrates, the depletion of “evolutionary powerhouses” in the tropics, and unpredictable emergent novelties. Despite this likelihood, we have only a rudimentary understanding of how we are altering the evolutionary future. As a result of our ignorance, conservation policies fail to reflect long-term evolutionary aspects of biodiversity loss.

Declines of biomes and biotas and the future of evolution

Although panel discussants disagreed whether the biodiversity crisis constitutes a mass extinction event, all agreed that current extinction rates are 50–500 times background and are increasing and that the consequences for the future evolution of life are serious. In response to the on-going rapid decline of biomes and homogenization of biotas, the panelists predicted changes in species geographic ranges, genetic risks of extinction, genetic assimilation, natural selection, mutation rates, the shortening of food chains, the increase in nutrient-enriched niches permitting the ascendancy of microbes, and the differential survival of ecological generalists. Rates of evolutionary processes will change in different groups, and speciation in the larger vertebrates is essentially over. Action taken over the next few decades will determine how impoverished the biosphere will be in 1,000 years when many species will suffer reduced evolvability and require interventionist genetic and ecological management. Whether the biota will continue to provide the dependable ecological services humans take for granted is less clear. The discussants offered recommendations, including two of paramount importance (concerning human populations and education), seven identifying specific scientific activities to better equip us for stewardship of the processes of evolution, and one suggesting that such stewardship is now our responsibility. The ultimate test of evolutionary biology as a science is not whether it solves the riddles of the past but rather whether it enables us to manage the future of the biosphere. Our inability to make clearer predictions about the future of evolution has serious consequences for both biodiversity and humanity.

Mirror self-recognition in the bottlenose dolphin: A case of cognitive convergence

The ability to recognize oneself in a mirror is an exceedingly rare capacity in the animal kingdom. To date, only humans and great apes have shown convincing evidence of mirror self-recognition. Two dolphins were exposed to reflective surfaces, and both demonstrated responses consistent with the use of the mirror to investigate marked parts of the body. This ability to use a mirror to inspect parts of the body is a striking example of evolutionary convergence with great apes and humans.

The challenge of contracting for technological information

Contracting to provide technological information (TI) is a significant challenge. TI is an unusual commodity in five ways. (i) TI is difficult to count and value; conventional indicators, such as patents and citations, hardly indicate value. TI is often sold at different prices to different parties. (ii) To value TI, it may be necessary to “give away the secret.” This danger, despite nondisclosure agreements, inhibits efforts to market TI. (iii) To prove its value, TI is often bundled into complete products, such as a computer chip or pharmaceutical product. Efficient exchange, by contrast, would involve merely the raw information. (iv) Sellers’ superior knowledge about TI’s value make buyers wary of overpaying. (v) Inefficient contracts are often designed to secure rents from TI. For example, licensing agreements charge more than marginal cost. These contracting difficulties affect the way TI is produced, encouraging self-reliance. This should be an advantage to large firms. However, small research and development firms spend more per employee than large firms, and nonprofit universities are major producers. Networks of organizational relationships, particularly between universities and industry, are critical in transmitting TI. Implicit barter—money for guidance—is common. Property rights for TI are hard to establish. Patents, quite suitable for better mousetraps, are inadequate for an era when we design better mice. Much TI is not patented, and what is patented sets fuzzy demarcations. New organizational forms are a promising approach to contracting difficulties for TI. Webs of relationships, formal and informal, involving universities, start-up firms, corporate giants, and venture capitalists play a major role in facilitating the production and spread of TI.

Genomic analysis of orthologous mouse and human olfactory receptor loci

Olfactory receptor (OR) genes represent ≈1% of genomic coding sequence in mammals, and these genes are clustered on multiple chromosomes in both the mouse and human genomes. We have taken a comparative genomics approach to identify features that may be involved in the dynamic evolution of this gene family and in the transcriptional control that results in a single OR gene expressed per olfactory neuron. We sequenced ≈350 kb of the murine P2 OR cluster and used synteny, gene linkage, and phylogenetic analysis to identify and sequence ≈111 kb of an orthologous cluster in the human genome. In total, 18 mouse and 8 human OR genes were identified, including 7 orthologs that appear to be functional in both species. Noncoding homology is evident between orthologs and generally is confined within the transcriptional unit. We find no evidence for common regulatory features shared among paralogs, and promoter regions generally do not contain strong promoter motifs. We discuss these observations, as well as OR clustering, in the context of evolutionary expansion and transcriptional regulation of OR repertoires.

General and specific brain regions involved in encoding and retrieval of events: what, where, and when.

Remembering an event involves not only what happened, but also where and when it occurred. We measured regional cerebral blood flow by positron emission tomography during initial encoding and subsequent retrieval of item, location, and time information. Multivariate image analysis showed that left frontal brain regions were always activated during encoding, and right superior frontal regions were always activated at retrieval. Pairwise image subtraction analyses revealed information-specific activations at (i) encoding, item information in left hippocampal, location information in right parietal, and time information in left fusiform regions; and (ii) retrieval, item in right inferior frontal and temporal, location in left frontal, and time in anterior cingulate cortices. These results point to the existence of general encoding and retrieval networks of episodic memory whose operations are augmented by unique brain areas recruited for processing specific aspects of remembered events.

Substitution rate calibration of small subunit ribosomal RNA identifies chlorarachniophyte endosymbionts as remnants of green algae.

Chlorarachniophytes are amoeboid algae with chlorophyll a and b containing plastids that are surrounded by four membranes instead of two as in plants and green algae. These extra membranes form important support for the hypothesis that chlorarachniophytes have acquired their plastids by the ingestion of another eukaryotic plastid-containing alga. Chlorarachniophytes also contain a small nucleus-like structure called the nucleomorph situated between the two inner and the two outer membranes surrounding the plastid. This nucleomorph is a remnant of the endosymbiont's nucleus and encodes, among other molecules, small subunit ribosomal RNA. Previous phylogenetic analyses on the basis of this molecule provided unexpected and contradictory evidence for the origin of the chlorarachniophyte endosymbiont. We developed a new method for measuring the substitution rates of the individual nucleotides of small subunit ribosomal RNA. From the resulting substitution rate distribution, we derived an equation that gives a more realistic relationship between sequence dissimilarity and evolutionary distance than equations previously available. Phylogenetic trees constructed on the basis of evolutionary distances computed by this new method clearly situate the chlorarachniophyte nucleomorphs among the green algae. Moreover, this relationship is confirmed by transversion analysis of the Chlorarachnion plastid small subunit ribosomal RNA.

Smoothness within ruggedness: the role of neutrality in adaptation.

RNA secondary structure folding algorithms predict the existence of connected networks of RNA sequences with identical structure. On such networks, evolving populations split into subpopulations, which diffuse independently in sequence space. This demands a distinction between two mutation thresholds: one at which genotypic information is lost and one at which phenotypic information is lost. In between, diffusion enables the search of vast areas in genotype space while still preserving the dominant phenotype. By this dynamic the success of phenotypic adaptation becomes much less sensitive to the initial conditions in genotype space.