Temporal variation, dormancy, and coexistence: A field test of the storage effect

Theoretical models suggest that overlapping generations, in combination with a temporally fluctuating environment, may allow the persistence of competitors that otherwise would not coexist. Despite extensive theoretical development, this “storage effect” hypothesis has received little empirical attention. Herein I present the first explicit mathematical analysis of the contribution of the storage effect to the dynamics of competing natural populations. In Oneida Lake, NY, data collected over the past 30 years show a striking negative correlation between the water-column densities of two species of suspension-feeding zooplankton, Daphnia galeata mendotae and Daphnia pulicaria. I have demonstrated competition between these two species and have shown that both possess long-lived eggs that establish overlapping generations. Moreover, recruitment to this long-lived stage varies annually, so that both daphnids have years in which they are favored (for recruitment) relative to their competitor. When the long-term population growth rates are calculated both with and without the effects of a variable environment, I show that D. galeata mendotae clearly cannot persist without the environmental variation and prolonged dormancy (i.e., storage effect) whereas D. pulicaria persists through consistently high per capita recruitment to the long-lived stage.

Spontaneous mutations recovered as mosaics in the mouse specific-locus test

The specific-locus test (SLT) detects new mutants among mice heterozygous for seven recessive visible markers. Spontaneous mutations can be manifested not only as singleton whole-body mutants in controls (for which we report new data), but as mosaics—either visible (manifesting mottled coat color) in the scored generation (G2) or masked, among the wild-type parental generation (G1). Masked G1 mosaics reveal themselves by producing clusters of whole-body mutants in G2. We provide evidence that most, if not all, mosaics detected in the SLT (both radiation and control progenies) result from a single-strand spontaneous mutation subsequent to the last premeiotic mitosis and before the first postmeiotic one of a parental genome—the “perigametic interval.” Such events in the genomes of the G1 and G0 result, respectively, in visible and masked 50:50 mosaics. Per cell cycle, the spontaneous mutation rate in the perigametic interval is much higher than that in pregamete mitotic divisions. A clearly different locus spectrum further supports the hypothesis of different origin, and casts further doubt on the validity of the doubling-dose risk-estimation method. Because mosaics cannot have arisen in mitotic germ cells, and are not induced by radiation exposure in the perigametic interval, they should not be included in calculations of radiation-induced germ-line mutation rates. For per-generation calculations, inclusion of mosaics yields a spontaneous frequency 1.7 times that calculated from singletons alone for mutations contributed by males; including both sexes, the multiple is 2.2.

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.

Estrogen alters behavior and forebrain c-fos expression in ovariectomized rats subjected to the forced swim test

Estrogen has been implicated in brain functions related to affective state, including hormone-related affective disorders in women. Although some reports suggest that estrogen appears to decrease vulnerability to affective disorders in certain cases, the mechanisms involved are unknown. We used the forced swim test (FST), a paradigm used to test the efficacy of antidepressants, and addressed the hypotheses that estrogen alters behavior of ovariectomized rats in the FST and the FST-induced expression of c-fos, a marker for neuronal activity, in the rat forebrain. The behaviors displayed included struggling, swimming, and immobility. One hour after the beginning of the test on day 2, the animals were perfused, and the brains were processed for c-fos immunocytochemistry. On day 1, the estradiol benzoate-treated animals spent significantly less time struggling and virtually no time in immobility and spent most of the time swimming. Control rats spent significantly more time struggling or being immobile during a comparable period. On day 2, similar behavioral patterns with still more pronounced differences were observed between estradiol benzoate and ovariectomized control groups in struggling, immobility, and swimming. Analysis of the mean number of c-fos immunoreactive cell nuclei showed a significant reduction in the estradiol benzoate versus control groups in areas of the forebrain relating to sensory, contextual, and integrative processing. Our results suggest that estrogen-induced neurochemical changes in forebrain neurons may translate into an altered behavioral output in the affective domain.

Conserved biological function between Pax-2 and Pax-5 in midbrain and cerebellum development: Evidence from targeted mutations

The development of two major subdivisions of the vertebrate nervous system, the midbrain and the cerebellum, is controlled by signals emanating from a constriction in the neural primordium called the midbrain/hindbrain organizer (Joyner, A. L. (1996) Trends Genet. 12, 15–201). The closely related transcription factors Pax-2 and Pax-5 exhibit an overlapping expression pattern very early in the developing midbrain/hindbrain junction. Experiments carried out in fish (Krauss, S., Maden, M., Holder, N. & Wilson, S. W. (1992) Nature (London) 360, 87–89) with neutralizing antibodies against Pax-b, the orthologue of Pax-2 in mouse, placed this gene family in an regulatory cascade necessary for the development of the midbrain and the cerebellum. The targeted mutation of Pax-5 has been reported to have only slight effects in the development of structures derived from the isthmic constriction, whereas the Pax-2 null mutant mice show a background-dependent phenotype with varying penetrance. To test a possible redundant function between Pax-2 and Pax-5 we analyzed the brain phenotypes of mice expressing different dosages of both genes. Our results demonstrate a conserved biological function of both proteins in midbrain/hindbrain regionalization. Additionally, we show that one allele of Pax-2, but not Pax-5, is necessary and sufficient for midbrain and cerebellum development in C57BL/6 mice.

DNA methylation is a reversible biological signal

The pattern of DNA methylation plays an important role in regulating different genome functions. To test the hypothesis that DNA methylation is a reversible biochemical process, we purified a DNA demethylase from human cells that catalyzes the cleavage of a methyl residue from 5-methyl cytosine and its release as methanol. We show that similar to DNA methyltransferase, DNA demethylase shows CpG dinucleotide specificity, can demethylate mdCpdG sites in different sequence contexts, and demethylates both fully methylated and hemimethylated DNA. Thus, contrary to the commonly accepted model, DNA methylation is a reversible signal, similar to other physiological biochemical modifications.

Structure-based assignment of the biochemical function of a hypothetical protein: A test case of structural genomics

Many small bacterial, archaebacterial, and eukaryotic genomes have been sequenced, and the larger eukaryotic genomes are predicted to be completely sequenced within the next decade. In all genomes sequenced to date, a large portion of these organisms’ predicted protein coding regions encode polypeptides of unknown biochemical, biophysical, and/or cellular functions. Three-dimensional structures of these proteins may suggest biochemical or biophysical functions. Here we report the crystal structure of one such protein, MJ0577, from a hyperthermophile, Methanococcus jannaschii, at 1.7-Å resolution. The structure contains a bound ATP, suggesting MJ0577 is an ATPase or an ATP-mediated molecular switch, which we confirm by biochemical experiments. Furthermore, the structure reveals different ATP binding motifs that are shared among many homologous hypothetical proteins in this family. This result indicates that structure-based assignment of molecular function is a viable approach for the large-scale biochemical assignment of proteins and for discovering new motifs, a basic premise of structural genomics.

An experimental test for lineage-specific position effects on alcohol dehydrogenase (Adh) genes in Drosophila

Independent transgene insertions differ in expression based on their location in the genome; these position effects are of interest because they reflect the influence of genome organization on gene regulation. Position effects also represent potentially insurmountable obstacles to the rigorous functional comparison of homologous genes from different species because (i) quantitative variation in expression of each gene across genomic positions (generalized position effects, or GPEs) may overwhelm differences between the genes of interest, or (ii) divergent genes may be differentially sensitive to position effects, reflecting unique interactions between each gene and its genomic milieu (lineage-specific position effects, or LSPEs). We have investigated both types of position-effect variation by applying our method of transgene coplacement, which allows comparisons of transgenes in the same position in the genome of Drosophila melanogaster. Here we report an experimental test for LSPE in Drosophila. The alcohol dehydrogenase (Adh) genes of D. melanogaster and Drosophila affinidisjuncta differ in both tissue distribution and amounts of ADH activity. Despite this striking regulatory divergence, we found a very high correlation in overall ADH activity between the genes of the two species when placed in the same genomic position as assayed in otherwise Adh-null adults and larvae. These results argue against the influence of LSPE for these sequences, although the effects of GPE are significant. Our new findings validate the coplacement approach and show that it greatly magnifies the power to detect differences in expression between transgenes. Transgene coplacement thus dramatically extends the range of functional and evolutionary questions that can be addressed by transgenic technology.

Verb generation in patients with focal frontal lesions: A neuropsychological test of neuroimaging findings

What are the neural bases of semantic memory? Traditional beliefs that the temporal lobes subserve the retrieval of semantic knowledge, arising from lesion studies, have been recently called into question by functional neuroimaging studies finding correlations between semantic retrieval and activity in left prefrontal cortex. Has neuroimaging taught us something new about the neural bases of cognition that older methods could not reveal or has it merely identified brain activity that is correlated with but not causally related to the process of semantic retrieval? We examined the ability of patients with focal frontal lesions to perform a task commonly used in neuroimaging experiments, the generation of semantically appropriate action words for concrete nouns, and found evidence of the necessity of the left inferior frontal gyrus for certain components of the verb generation task. Notably, these components did not include semantic retrieval per se.

Declining brain activity in cognitively normal apolipoprotein E ɛ4 heterozygotes: A foundation for using positron emission tomography to efficiently test treatments to prevent Alzheimer's disease

Cross-sectional positron emission tomography (PET) studies find that cognitively normal carriers of the apolipoprotein E (APOE) ɛ4 allele, a common Alzheimer's susceptibility gene, have abnormally low measurements of the cerebral metabolic rate for glucose (CMRgl) in the same regions as patients with Alzheimer's dementia. In this article, we characterize longitudinal CMRgl declines in cognitively normal ɛ4 heterozygotes, estimate the power of PET to test the efficacy of treatments to attenuate these declines in 2 years, and consider how this paradigm could be used to efficiently test the potential of candidate therapies for the prevention of Alzheimer's disease. We studied 10 cognitively normal ɛ4 heterozygotes and 15 ɛ4 noncarriers 50–63 years of age with a reported family history of Alzheimer's dementia before and after an interval of approximately 2 years. The ɛ4 heterozygotes had significant CMRgl declines in the vicinity of temporal, posterior cingulate, and prefrontal cortex, basal forebrain, parahippocampal gyrus, and thalamus, and these declines were significantly greater than those in the ɛ4 noncarriers. In testing candidate primary prevention therapies, we estimate that between 50 and 115 cognitively normal ɛ4 heterozygotes are needed per active and placebo treatment group to detect a 25% attenuation in these CMRgl declines with 80% power and P = 0.005 in 2 years. Assuming these CMRgl declines are related to the predisposition to Alzheimer's dementia, this study provides a paradigm for testing the potential of treatments to prevent the disorder without having to study thousands of research subjects or wait many years to determine whether or when treated individuals develop symptoms.

RNA virus error catastrophe: Direct molecular test by using ribavirin

RNA viruses evolve rapidly. One source of this ability to rapidly change is the apparently high mutation frequency in RNA virus populations. A high mutation frequency is a central tenet of the quasispecies theory. A corollary of the quasispecies theory postulates that, given their high mutation frequency, animal RNA viruses may be susceptible to error catastrophe, where they undergo a sharp drop in viability after a modest increase in mutation frequency. We recently showed that the important broad-spectrum antiviral drug ribavirin (currently used to treat hepatitis C virus infections, among others) is an RNA virus mutagen, and we proposed that ribavirin's antiviral effect is by forcing RNA viruses into error catastrophe. However, a direct demonstration of error catastrophe has not been made for ribavirin or any RNA virus mutagen. Here we describe a direct demonstration of error catastrophe by using ribavirin as the mutagen and poliovirus as a model RNA virus. We demonstrate that ribavirin's antiviral activity is exerted directly through lethal mutagenesis of the viral genetic material. A 99.3% loss in viral genome infectivity is observed after a single round of virus infection in ribavirin concentrations sufficient to cause a 9.7-fold increase in mutagenesis. Compiling data on both the mutation levels and the specific infectivities of poliovirus genomes produced in the presence of ribavirin, we have constructed a graph of error catastrophe showing that normal poliovirus indeed exists at the edge of viability. These data suggest that RNA virus mutagens may represent a promising new class of antiviral drugs.