Drosophila melanogaster does not share the telomeric repeat sequence of another invertebrate, Ascaris lumbricoides

Summary The DNA at the chromosomal termini of all eukaryotes from which it has been isolated contains a characteristic sequence motif consisting of tandem arrays of a regular or irregular repeat unit. These terminal repeats are thought to be essential for the maintenance of the chromosome ends. The sequences of the terminal repeats of all vertebrates studied thus far are identical and are similar enough to those of higher plants and some protozoans to cross-hybridize. However, previous studies have not detected cross-hybridization between the DNA of Drosophila mélanogaster and the terminal DNA sequences of any of several organisms tested. Recently, the first terminal DNA clone from a multicellular invertebrate, that of Ascaris lumbricoides, was reported also to consist of a tandem reiteration of a short sequence similar to those previously identified for other eukaryotes. Here I show that a probe for this sequence from A. lumbricoides fails to hybridize delectably to the DNA of D. melanogaster. Thus, in contrast to their conservation among vertebrates, the terminal chromosomal sequences appear not to be shared by all metazoan invertebrates.     

Molecular correlates of reproductive isolation

Evolution of reproductive isolation as a byproduct of genetic divergence in isolated populations is the dominant (albeit not exclusive) mode of speciation in sexual animals. But little is known about the factors linking speciation to general divergence. Several authors have argued that allopatric speciation should proceed more rapidly if isolated populations also experience divergent selection. Reproductive isolation between allopatric populations is not subject to direct selection; it can accumulate only by random drift or as a fortuitous byproduct of selection on other traits. Here I present a novel analysis of published data, demonstrating that pre- and postmating isolation of Drosophila species are more tightly correlated with allozyme divergence than with silent DNA divergence. Inasmuch as proteins are more subject to the action of natural selection than are silent DNA polymorphisms, this result provides broad support for a model of selection-mediated allopatric speciation.     

Divergent selection and phenotypic plasticity during incipient speciation in Lake Victoria cichlid fish

Divergent selection acting on several different traits that cause multidimensional shifts are supposed to promote speciation, but the outcome of this process is highly dependent on the balance between the strength of selection vs. gene flow. Here, we studied a pair of sister species of Lake Victoria cichlids at a location where they hybridize and tested the hypothesis that divergent selection acting on several traits can maintain phenotypic differentiation despite gene flow. To explore the possible role of selection we tested for correlations between phenotypes and environment and compared phenotypic divergence (P_ST) with that based on neutral markers (F_ST). We found indications for disruptive selection acting on male breeding colour and divergent selection acting on several morphological traits. By performing common garden experiments we also separated the environmental and heritable components of divergence and found evidence for phenotypic plasticity in some morphological traits contributing to species differences.     

Sequences isolated from aDrosophila early-ecdysone puff are expressed in rat liver

We have studied the presence of a cloned fragment of DNA from Drosophila melanogaster in other organisms by means of nucleic acid hybridization analysis. The isolated region is localized in polytene chromosomes at the 63F subdivision. This region includes a puff that responds within minutes to ecdysone stimulation. We have found that 63F DNA from D. melanogaster hybridizes 'in situ' to both DNA and RNA from D. simulans, D. teissieri, and D. hydei. In all these species the isolated DNA remains associated with one early-ecdysone stimulated puff. The isolated Drosophila recombinant DNA is also complementary to polyadenylated RNA from foetal and adult rat liver but fails to hybridize to the nonpolyadenylated RNA classes from both sources and to polyadenylated RNA from rat mammary glands.     

HALDANE'S RULE AND SEX BIASSED GENE FLOW BETWEEN TWO HYBRIDIZING FLYCATCHER SPECIES (FICEDULA ALBICOLLIS AND F. HYPOLEUCA,AVES: MUSCICAPIDAE)

The collared flycatcher (Ficedula albicollis) and the pied flycatcher (F. hypoleuca) hybridize where their geographic ranges overlap. Restriction fragment comparison of 5% of the mitochondrial genome showed a sequence divergence of 10% between these flycatcher species. This degree of sequence divergence between a closely related pair of bird species is unusually high and contrasts with the low level of divergence between F. albicollis and F. hypoleuca in nuclear genes (Nei's D = 0.0006) revealed by enzyme electrophoresis. The low nuclear differentiation is explained by sex biassed gene flow and introgression in nuclear genes (via fertile male hybrids), while the high mitochondrial DNA sequence divergence is preserved by sterility of female hybrids, which prevents mitochondrial introgression. This pattern is in accordance with Haldane's rule and is supported by field data on hybrid fertility. The high mtDNA differentiation could be explained by transfer of mitochondrial DNA from a third species during a past period of hybridization.     

Divergence of a speciation trait through artificial selection: Insights into constraints,by-product effects and sexual isolation

Speciation and sexual isolation often occur when divergent female mating preferences target male secondary sexual traits. Despite the importance of such male signals, little is known about their evolvability and genetic linkage to other traits during speciation. To answer these questions, we imposed divergent artificial selection for 10 non-overlapping generations on the Inter-Pulse-Interval (IPI) of male courtship songs; which has been previously shown to be a major species recognition trait for females in the Drosophila athabasca species complex. Focusing on one of the species, Drosophila mahican (previously known as EA race), we examined IPI's: (1) rate of divergence, (2) response to selection in different directions, (3) genetic architecture of divergence and (4) by-product effects on other traits that have diverged in the species complex. We found rapid and consistent response for higher IPI but less response to lower IPI; implying asymmetrical constraints. Genetic divergence in IPI differed from natural species in X versus autosome contribution and in dominance, suggesting that evolution may take different paths. Finally, selection on IPI did not alter other components of male songs, or other ecological traits, and did not cause divergence in female preferences, as evidenced by lack of sexual isolation. This suggests that divergence of male courtship song IPI is unconstrained by genetic linkage with other traits in this system. This lack of linkage between male signals and other traits implies that female preferences or ecological selection can co-opt and mould specific male signals for species recognition free of genetic constraints from other traits.     

Geographic partitioning of chloroplast DNA variation in the genusDatisca (Datiscaceae)

Datisca (Datiscaceae) is a ditypic genus with an intercontinentally disjunct distribution. Chloroplast DNA restriction site data was obtained from 23 populations and four 10–20 year old herbarium specimens ofD. glomerata and three populations ofD. cannabina from throughout their geographic ranges in western North America and southwest-central Asia, respectively. InD. glomerata, plastome diversity is partitioned geographically. All populations from southern California have a common plastome, while most populations north of this region share a relatively divergent plastome (0.49% sequence divergence). Likewise, these plastomes are highly divergent (0.87% mean sequence divergence) from those found inD. cannabina. Biogeographic processes dating to the Pleistocene and Late Miocene may be responsible for these intra- and interspecific patterns of chloroplast DNA divergence.     

Hybridization between genetically and morphologically divergent forms of Rhagada (Gastropoda: Camaenidae) snails at a zone of secondary contact

Congeneric sympatry is rare in Australian camaenid land snails, and the reasons are poorly understood. As the first example of contact between species of the Western Australian genus Rhagada, we examined the parapatric zone between the coastally distributed Rhagada convicta and an undescribed inland species, which differ in size, shape, and banding of their shells and in their reproductive systems. Phylogenetic analysis of sequences of the mitochondrial 16S RNA gene indicated secondary contact between two distinct clades, with an average sequence divergence of 6.5%. Despite these substantial differences, the two forms hybridize in the narrow contact zone, resulting in intermediate phenotypes for shells and reproductive anatomy, and incongruence of the mitochondrial DNA (mtDNA) lineage and morphology. Hybridization appears to be asymmetric, however, as all morphological intermediates possessed the R. convicta mtDNA. Interbreeding between these deeply divergent forms challenges the use of reproductive structures as indicators of reproductive isolation in snail taxonomy. This is the first contact zone detected between mainland species of Rhagada and indicates that the widespread pattern of geographical replacement is not simply a result of allopatric divergence without subsequent contact. Instead, the perspective for understanding the radiation and geographical distributions must include interactions between the species. © 2014 The Linnean Society of London, Biological Journal of the Linnean Society, 2015, 114 , 348–362.     

EVOLUTIONARY IMPLICATIONS OF DNA DIVERGENCE IN THE DROSOPHILA OBSCURA GROUP

Using DNA–DNA hybridization, we have determined the degree of single-copy DNA (scDNA) divergence among eight species of the Drosophila obscura group. These include Old World and New World species as well as members of two subgroups. Contrary to classical systematics, members of the affinis subgroup are more closely related to American members of the obscura subgroup than are Old World species. The Old World species are not a monophyletic group. The degree of scDNA divergence among species is not necessarily correlated with morphology, chromosomal divergence, or ability to form hybrids. A unique pattern of hybrid formation was found: species separated by a ΔT_m of 6.5°C can form hybrids whereas species separated by a ΔT_m of 2.5°C cannot. As with other groups of Drosophila, the obscura group has discrete parts of the genome evolving at very different rates. The slow evolving fraction of the nuclear genome is evolving at about the same rate as mitochondrial DNA. The additional scDNA divergence accompanying the step from partial reproductive isolation (between North American pseudoobscura and the isolated Bogotà population) to full isolation is very small. The resolution of the technique was challenged by five closely related taxa with a maximum ΔT_m of 2.5°C separating them; the taxa were unambiguously resolved and the “correct” phylogeny recovered. Finally, there is some indication that scDNA in the obscura group may be evolving considerably slower than in the melanogaster subgroup.     

Evolutionary distances in hawaiian drosophila measured by DNA reassociation

Summary Comparisons of the sequence divergence of three species of Hawaiian Drosophila have been made by hybridization of single-copy tracer DNA of each of the species with driver DNA from each species, and measurement of the average melting temperature (Tma) in a chaotropic solvent (2.4 M tetraethylammonium chloride) which minimizes differences due to base composition. Correction was made for the length of hybrid duplex regions to obtain the reduction in thermal stability due to divergence.An accuracy of ± 0.2°C was achieved and the mean reduction in Tm for hybridization betweenD. heteroneura andD. silvestris (found only on the island of Hawaii) was 0.55°C and betweenD. picticornis, found only on the island of Kauai, and the other two species was 2.13°C. The rate of DNA change is estimated to be between 0.2 and 0.4%/My by assuming that theD. heteroneura-D. silvestris divergence occurred 0.8 My ago and the divergence between these species andD. picticornis between 4 and 6 My ago.The general single copy DNA sequence divergence appears to be very much greater than the minimal coding region sequence divergence previously estimated from allozyme studies.     

Use of a cellulase-encoding gene probe to reveal restriction fragment length polymorphisms among ruminal strains ofBacteroides succinogenes

A cloned DNA fragment specifying an endoglucanase fromBacteroides succinogenes strain BL2 was shown to hybridize under nonstringent conditions to different BamH1 fragments of chromosomal DNA from each of five rumen strains ofB. succinogenes. Direct binding of BL2 total chromosomal DNA to DNA from the other four strains was between 16% and 42% of homologous binding, confirming a high degree of interstrain divergence.Bacteroides succinogenes BL2 chromosomal DNA did not show detectable hybridization with DNA from any of 15 strains of 11 other species of rumen bacteria, in tests carried out with NaOH-lysed cells on filters.     

Evolution of single-copy DNA and the ADH gene in seven drosophilids

Summary Single-copy DNA was isolated fromDrosophila melanogaster and hybridized with total genomic DNA ofD. melanogaster, D. mauritiana, D. simulans, D. pseudoobscura, D. willistoni, D. hydei andD. virilis. The duplexes were thermally eluted from hydroxyapatite and the data used to assess the relatedness of each species toD. melanogaster. The general pattern of relatedness was similar to that predicted by morphological methods but with some notable exceptions. The rate of nucleotide substitution was estimated to be greater than 0.66% of bases per million years. An unexpected, rapidly evolving component ofD. melanogaster single-copy DNA was identified. The relatedness of these species was also studied with respect to the gene coding for alcohol dehydrogenase (ADH). The ADH gene, previously cloned fromD. melanogaster (Goldberg 1980), was hybridized with Southern blots of genomic digests of the seven species. The intensity and position of the hybridizing bands suggest the amount of divergence of the gene. Divergence was quantitated by reassociation of a fragment of the cloned ADH gene with total DNA of the seven drosophilids and thermal elution of the resultant duplexes from hydroxyapatite. The ADH gene was isolated from genomic clone libraries ofD. melanogaster, D. simulans andD. mauritiana and further studied by comparison of position of restriction sites. Species relationships deduced from comparison of total single-copy DNA and the ADH gene were consistent, demonstrating that a single gene can reflect divergence of the entire genome.     

Wolbachia infection decreased the resistance of Drosophila to lead

Background

The heavy metal lead has been shown to be associated with a genotoxic risk. Drosophila melanogaster is a model organism commonly utilized in genetic toxicology testing. The endosymbionts — Wolbachia are now very common in both wild populations and laboratory stocks of Drosophila. Wolbachia may induce resistance to pathogenic viruses, filarial nematodes and Plasmodium in fruit fly and mosquito hosts. However the effect of Wolbachia infection on the resistance of their hosts to heavy metal is unknown.

Methodology/Principal Findings

Manipulating the lead content in the diet of Drosophila melanogaster, we found that lead consumption had no different effects on developmental time between Wolbachia-infected (Dmel wMel) and –uninfected (Dmel T) flies. While in Pb-contaminated medium, significantly reduced amount of pupae and adults of Dmel wMel were emerged, and Dmel wMel adults had significantly shorter longevity than that of Dmel T flies. Lead infusion in diet resulted in significantly decreased superoxide dismutase (SOD) activity in Dmel T flies (P<0.05), but not in Dmel wMel flies. Correspondingly, lead cultures induced a 10.8 fold increase in malonaldehyde (MDA) contents in Dmel T larvae (P<0.05). While in Dmel wMel larvae, it resulted in only a 1.3 fold increase. By quantitative RT-PCR, we showed that lead infused medium caused significantly increased expression level of relish and CecA2 genes in Dmel T flies (P<0.01). Lead cultures did not change dramatically the expression of these genes in Dmel wMel flies.

Conclusions/Significance

These results suggest that Wolbachia infection decreased the resistance of Drosophila to lead likely by limiting the production of peroxides resulted from lead, thus being unable to activate the immunological pathway in the host to prevent them from lead damage. This represents a novel Wolbachia–host interaction and provides information that researchers working on Drosophila toxicology should take in consideration the presence of Wolbachia in the stocks they are analyzing.     

PARALLEL EVOLUTION OF LAKE-STREAM PAIRS OF THREESPINE STICKLEBACKS (GASTEROSTEUS) INFERRED FROM MITOCHONDRIAL DNA VARIATION

Three drainage systems in British Columbia, Canada, contain divergent parapatric lake-stream pairs of threespine sticklebacks (Gasterosteus aculeatus): Drizzle and Mayer Lakes on Graham Island, Queen Charlotte Islands, and Misty Lake on northeastern Vancouver Island. Ecological and morphological differences between members of all three lake-stream pairs are strikingly similar; lake fish are melanistic and slim bodied with smaller mouths and more gill rakers than the mottled-brown and robust-bodied stream sticklebacks. We estimated the level of genetic divergence between lake and stream fish in Misty Lake and tested hypotheses of single versus multiple origins of the pairs by assaying mitochondrial DNA (mtDNA) restriction site variation in samples from the three lake systems. MtDNA analysis revealed the existence of two highly divergent lineages differing by 2.7% in sequence. One lineage predominated in Misty stream fish (73%), whereas the other lineage predominated in Misty Lake samples (96%). Comparable forms (lake or stream) in the different lakes did not cluster together in terms of mtDNA nucleotide divergence, suggesting that the pairs have had independent origins. We concluded that: (1) divergent mtDNA lineages in North Pacific sticklebacks stem from historical isolation in the two major glacial refugia proposed for the North Pacific (Beringia and Cascadia); (2) the stream and lake pair in Misty Lake are distinct gene pools; (3) the divergence between parapatric lake and stream Gasterosteus represents parallel evolution having occurred at least twice in the North Pacific; and (4) different scales of evolutionary divergence exist in North Pacific Gasterosteus, that is, a relatively ancient divergence of mtDNA clades as well as recent (i.e., postglacial) divergence of ecotypes within major clades.     

Genomic divergence between Spanish Littorina saxatilis ecotypes unravels limited admixture and extensive parallelism associated with population history

The rough periwinkle, Littorina saxatilis, is a model system for studying parallel ecological speciation in microparapatry. Phenotypically parallel wave‐adapted and crab‐adapted ecotypes that hybridize within the middle shore are replicated along the northwestern coast of Spain and have likely arisen from two separate glacial refugia. We tested whether greater geographic separation corresponding to reduced opportunity for contemporary or historical gene flow between parallel ecotypes resulted in less parallel genomic divergence. We sequenced double‐digested restriction‐associated DNA (ddRAD) libraries from individual snails from upper, mid, and low intertidal levels of three separate sites colonized from two separate refugia. Outlier analysis of 4256 SNP markers identified 34.4% sharing of divergent loci between two geographically close sites; however, these sites each shared only 9.9%–15.1% of their divergent loci with a third more‐distant site. STRUCTURE analysis revealed that genotypes from only three of 166 phenotypically intermediate mid‐shore individuals appeared to result from recent hybridization, suggesting that hybrids cannot be reliably identified using shell traits. Hierarchical AMOVA indicated that the primary source of genomic differentiation was geographic separation, but also revealed greater similarity of the same ecotype across the two geographically close sites than previously estimated with dominant markers. These results from a model system for ecological speciation suggest that genomic parallelism is affected by the opportunity for historical or contemporary gene flow between populations.     

Sequence and secondary structure of the central domain ofDrosophila 26S rRNA: A universal model for the central domain of the large rRNA containing the region in which the central break may happen

Summary An 890-bp sequence from the central region ofDrosophila melanogaster 26S ribosomal DNA (rDNA) has been determined and used in an extensive comparative analysis of the central domain of the large subunit ribosomal RNA (lrRNA) from prokaryotes, organelles, and eukaryotes. An alignment of these different sequences has allowed us to precisely map the regions of the central domain that have highly diverged during evolution. Using this sequence comparison, we have derived a secondary structure model of the central domain ofDrosophila 26S ribosomal RNA (rRNA). We show that a large part of this model can be applied to the central domain of lrRNA from prokaryotes, eukaryotes, and organelles, therefore defining a universal common structural core. Likewise, a comparative study of the secondary structure of the divergent regions has been performed in several organisms. The results show that, despite a nearly complete divergence in their length and sequence, a common structural core is also present in divergent regions. In some organisms, one or two of the divergent regions of the central domain are removed by processing events. The sequence and structure of these regions (fragmentation spacers) have been compared to those of the corresponding divergent regions that remain part of the mature rRNA in other species.     

Rates of DNA Evolution in Drosophila Depend on Function and Developmental Stage of Expression

DNA-sequence divergence of genes expressed in the embryonic stage was compared with the divergence of genes expressed in adults for 13 species of Drosophila representing various degrees of relatedness. DNA-DNA hybridization experiments were conducted using as tracers complementary DNA (cDNA) reversed transcribed from poly(A)(+) mRNA isolated from different developmental stages. The results indicate: (1) cDNA is less diverged than total single-copy DNA; (2) cDNA sequences are not in the rapidly evolving fraction of the single-copy genome of Drosophila; (3) early in evolutionary divergence embryonic messages are about half as diverged as adult messages; sequence data from some of the species compared indicate this is likely due to differences in rates of silent substitutions in genes expressed at different stages of development; and (4) at greater evolutionary distance, the differences in embryonic and adult messages disappear; this could be due to lineage-specific shifts in codon usage.     

Cellular senescence involves stochastic processes causing loss of expression of differentiated function genes: visualization by in situ hybridization for steroid 17 alpha-hydroxylase in bovine adrenocortical cells

When grown for long periods in culture, bovine adrenocortical cells lose the expression of a differentiated function gene, steroid 17 alpha-hydroxylase. Previously, we documented a decline in 17 alpha-hydroxylase mRNA with increasing culture passage level after induction with cyclic AMP (P. J. Hornsby et al., 1987, Proc. Natl. Acad. Sci. USA 84, 1580). We used in situ hybridization to investigate the loss of expression of this gene during cellular senescence at an individual cell level. In primary cultures, cells were uniformly positive for hybridization with cDNA for 17 alpha-hydroxylase after cyclic AMP induction. After two passages, cultures comprised a mixture of hybridizing and nonhybridizing cells. Cells appeared either to hybridize at a level comparable to that in primary cultures or to be nonhybridizing. When in situ hybridization was combined with immunofluorescence, cells positive for immunofluorescence were also positive for hybridization. Senescing mass cultures showed decreasing numbers of positive cells, and after 30 passages cultures comprised entirely nonhybridizing cells. Thus, the previously observed decline in overall 17 alpha-hydroxylase mRNA levels results from a decline in the fraction of expressing cells in the culture, and the rate of loss of expressing cells is in agreement with the rate of loss of total 17 alpha-hydroxylase mRNA. Primary clones, even when isolated at an early stage of clonal expansion, had mixtures of subclones of hybridizing and nonhybridizing cells. On recloning, hybridizing subclones usually produced uniformly nonhybridizing sub-subclones. Some subclones within primary clones had a morphology associated with replicative senescence (flattened cells with sparse intercellular contacts), yet had high numbers of hybridizing cells. We conclude that, in both mass and clonal populations, cells initially expressing 17 alpha-hydroxylase rapidly give rise to clones of nonexpressing cells. Such cells are continually derived by a stochastic process from cells originally expressing the gene.