Comparative sequence analysis and patterns of covariation in RNA secondary structures.

A novel method of RNA secondary structure prediction based on a comparison of nucleotide sequences is described. This method correctly predicts nearly all evolutionarily conserved secondary structures of five different RNAs: tRNA, 5S rRNA, bacterial ribonuclease P (RNase P) RNA, eukaryotic small subunit rRNA, and the 3' untranslated region (UTR) of the Drosophila bicoid (bcd) mRNA. Furthermore, covariations occurring in the helices of these conserved RNA structures are analyzed. Two physical parameters are found to be important determinants of the evolution of compensatory mutations: the length of a helix and the distance between base-pairing nucleotides. For the helices of bcd 3' UTR mRNA and RNase P RNA, a positive correlation between the rate of compensatory evolution and helix length is found. The analysis of Drosophila bcd 3' UTR mRNA further revealed that the rate of compensatory evolution decreases with the physical distance between base-pairing residues. This result is in qualitative agreement with Kimura's model of compensatory fitness interactions, which assumes that mutations occurring in RNA helices are individually deleterious but become neutral in appropriate combinations.

Linkage disequilibria and the site frequency spectra in the su(s) and su(w(a)) regions of the Drosophila melanogaster X chromosome.

Over the last decade, surveys of DNA sequence variation in natural populations of several Drosophila species and other taxa have established that polymorphism is reduced in genomic regions characterized by low rates of crossing over per physical length. Parallel studies have also established that divergence between species is not reduced in these same genomic regions, thus eliminating explanations that rely on a correlation between the rates of mutation and crossing over. Several theoretical models (directional hitchhiking, background selection, and random environment) have been proposed as population genetic explanations. In this study samples from an African population (n = 50) and a European population (n = 51) were surveyed at the su(s) (1955 bp) and su(w(a)) (3213 bp) loci for DNA sequence polymorphism, utilizing a stratified SSCP/DNA sequencing protocol. These loci are located near the telomere of the X chromosome, in a region of reduced crossing over per physical length, and exhibit a significant reduction in DNA sequence polymorphism. Unlike most previously surveyed, these loci reveal substantial skews toward rare site frequencies, consistent with the predictions of directional hitchhiking and random environment models and inconsistent with the general predictions of the background selection model (or neutral theory). No evidence for excess geographic differentiation at these loci is observed. Although linkage disequilibrium is observed between closely linked sites within these loci, many recombination events in the genealogy of the sampled alleles can be inferred and the genomic scale of linkage disequilibrium, measured in base pairs between sites, is the same as that observed for loci in regions of normal crossing over. We conclude that gene conversion must be high in these regions of low crossing over.

The hitchhiking effect on the site frequency spectrum of DNA polymorphisms.

The level of DNA sequence variation is reduced in regions of the Drosophila melanogaster genome where the rate of crossing over per physical distance is also reduced. This observation has been interpreted as support for the simple model of genetic hitchhiking, in which directional selection on rare variants, e.g., newly arising advantageous mutants, sweeps linked neutral alleles to fixation, thus eliminating polymorphisms near the selected site. However, the frequency spectra of segregating sites of several loci from some populations exhibiting reduced levels of nucleotide diversity and reduced numbers of segregating sites did not appear different from what would be expected under a neutral equilibrium model. Specifically, a skew toward an excess of rare sites was not observed in these samples, as measured by Tajima's D. Because this skew was predicted by a simple hitchhiking model, yet it had never been expressed quantitatively and compared directly to DNA polymorphism data, this paper investigates the hitchhiking effect on the site frequency spectrum, as measured by Tajima's D and several other statistics, using a computer simulation model based on the coalescent process and recurrent hitchhiking events. The results presented here demonstrate that under the simple hitchhiking model (1) the expected value of Tajima's D is large and negative (indicating a skew toward rare variants), (2) that Tajima's test has reasonable power to detect a skew in the frequency spectrum for parameters comparable to those from actual data sets, and (3) that the Tajima's Ds observed in several data sets are very unlikely to have been the result of simple hitchhiking. Consequently, the simple hitchhiking model is not a sufficient explanation for the DNA polymorphism at those loci exhibiting a decreased number of segregating sites yet not exhibiting a skew in the frequency spectrum.

RNA secondary structure and compensatory evolution.

The classic concept of epistatic fitness interactions between genes has been extended to study interactions within gene regions, especially between nucleotides that are important in maintaining pre-mRNA/mRNA secondary structures. It is shown that the majority of linkage disequilibria found within the Drosophila Adh gene are likely to be caused by epistatic selection operating on RNA secondary structures. A recently proposed method of RNA secondary structure prediction based on DNA sequence comparisons is reviewed and applied to several types of RNAs, including tRNA, rRNA, and mRNA. The patterns of covariation in these RNAs are analyzed based on Kimura's compensatory evolution model. The results suggest that this model describes the substitution process in the pairing regions (helices) of RNA secondary structures well when the helices are evolutionarily conserved and thermodynamically stable, but fails in some other cases. Epistatic selection maintaining pre-mRNA/mRNA secondary structures is compared to weak selective forces that determine features such as base composition and synonymous codon usage. The relationships among these forces and their relative strengths are addressed. Finally, our mutagenesis experiments using the Drosophila Adh locus are reviewed. These experiments analyze long-range compensatory interactions between the 5' and 3' ends of Adh mRNA, the different constraints on secondary structures in introns and exons, and the possible role of secondary structures in RNA splicing.

Loss of a paternal chromosome causes developmental anomalies among Drosophila hybrids.

Hybrids between Drosophila virilis and D. lummei suffer from developmental anomalies. Previous reports also suggest that these hybrids lose the D. lummei sixth chromosome early in development. Genetic and cytological data presented here confirm the loss of the microchromosome from both the soma and the germ-line of these hybrids and provide strong evidence that this loss causes the hybrid developmental anomalies.

A test of the background selection hypothesis based on nucleotide data from Drosophila ananassae.

We estimated DNA sequence variation within and between four populations of Drosophila ananassae at Om(1D) and vermilion (v) by using single-strand conformation polymorphism analysis and direct DNA sequencing. Om(1D) is located on the X chromosome in a region with a normal recombination rate; v is in a region of low recombination. In each population, levels of nucleotide diversity at v are reduced 10- to 25-fold relative to those at Om(1D). Divergence between D. ananassae and its sibling species D. pallidosa, however, is comparable for both loci. This lack of correlation between levels of polymorphism and divergence led to the rejection of a constant-rate, neutral model. To distinguish among alternative models, we propose a test of the background selection hypothesis based on the observed pattern of differentiation between populations. Although the degree of differentiation (measured by FST) among all pairs of subpopulations is similar at Om(1D), we found substantial differences at v. The two northern populations from Burma and Nepal are very homogeneous, whereas comparisons between northern and southern populations (e.g., between Nepal and middle India) produced large FST values. A coalescent-based simulation of the background selection model (in a geographically structured species with a finite number of demes) showed that the observed homogeneity among the northern populations is inconsistent with the background selection hypothesis. Instead, it may have been caused by a recent hitchhiking event that was limited to the northern species range.

A population-based serosurveillance of syphilis in Costa Rica.

As part of a case-control study to investigate the high incidence of cervical cancer in Costa Rican women, the seroprevalence of the treponematoses, in particular, syphilis was determined. In each age group, women with a history of two or more sex partners were two to four times more likely to be seroreactive in tests for syphilis than women with zero or one sex partner. The highest percentage of reactive results in the microhemagglutination assay for antibodies to Treponema pallidum (MHA-TP) was seen in samples from women aged 50-59 who had had two or more lifetime partners (23.8%). Three observations from our study support reactivity due to syphilis rather than yaws or pinta: (1) a similar percent of reactive rapid plasma reagin (RPR) card test results among MHA-TP reactors in the two age groups of women who were surveyed (42 vs. 49%) was observed; (2) women who were seroreactive in the MHA-TP had multiple risk factors for STD [low socioeconomic status (9.4%), urban residence (22.8%), first intercourse under 16 years of age (14.1%), and multiple sex partners (26.3%)], and (3) only sexually experienced women had reactive results in the MHA-TP test.