Rumi Kondo

Man's place in hominoidea revealed by mitochondrial DNA genealogy

SummaryMolecular biology has resurrected C. Darwin and T.H. Huxleys question about the origin of humans, but the precise branching pattern and dating remain controversial. To settle this issue, a large amount of sequence information is required. We determined mitochondrial (mt) DNA sequences for five hominoids; pygmy and common chimpanzees, gorilla, orangutan, and siamang. The common region compared with the known human sequence is 4759 by long, encompassing genes for 11 transfer RNAs and 6 proteins. Because of the high substitution rates in mammalian mtDNA and an unprecedentedly large region compared, the sequence differences clearly indicate that the closest relatives to human are chimpanzees rather than gorilla. For dating the divergences of human, chimpanzee, and gorilla, we used only unsaturated parts of sequence differences in which the mtDNA genealogy is not obscured by multiple substitutions. The result suggests that gorilla branched off 7.7 ± 0.7 million years (Myr) ago and human 4.7 ± 0.5 Myr ago; the time difference between these divergences being as long as 3 Myr.

Evolution of hominoid mitochondrial DNA with special reference to the silent substitution rate over the genome

SummaryFocusing on the synonymous substitution rate, we carried out detailed sequence analyses of hominoid mitochondrial (mt) DNAs of ca. 5-kb length. Owing to the outnumbered transitions and strong biases in the base compositions, synonymous substitutions in mtDNA reach rapidly a rather low saturation level. The extent of the compositional biases differs from gene to gene. Such changes in base compositions, even if small, can bring about considerable variation in observed synonymous differences and may result in the region-dependent estimate of the synonymous substitution rate. We demonstrate that such a region dependency is due to a failure to take proper account of heterogeneous compositional biases from gene to gene but that the actual synonymous substitution rate is rather uniform. The synonymous substitution rate thus estimated is 2.37 ± 0.11 × 10−8 per site per year and comparable to the overall rate for the noncoding region. On the other hand, the rate of nonsynonymous substitutions differs considerably from gene to gene, as expected under the neutral theory of molecular evolution. The lowest rate is 0.8 × 10−9 per site per year forCOI and the highest rate is 4.5 × 10−9 forATPase 8, the degree of functional constraints (measured by the ratio of the nonsynonymous to the synonymous substitution rate) being 0.03 and 0.19, respectively. Transfer RNA (tRNA) genes also show variability in the base contents and thus in the nucleotide differences. The average rate for 11 tRNAs contained in the 5-kb region is 3.9 × 10−9 per site per year. The nucleotide substitutions in the genome suggest that the transition rate is about 17 times faster than the transversion rate.

Diversification of olfactory receptor genes in the Japanese medaka fish, Oryzias latipes

Vertebrate olfactory receptors (OR) exists as the largest multigene family, scattered throughout the genome in clusters. Studies have shown that different animals possess remarkably diverse set of OR genes to recognize diverse odor molecules. In order to examine the evolutionary process of OR diversification, we examined three OR gene subfamilies from Japanese medaka fish (seven lines sampled from four populations). For each subfamily, the sequences of ancestral genes were inferred based on distance method. Examination of d(N)/d(S) ratios for each branch of phylogenetic trees suggested that purifying selection is the major force of evolution in medaka OR genes. However, for the mfOR1 and mfOR2 paralogous gene pairs, a nonrandom distribution of fixed amino acid changes and the d(N)>d(S) in a branch suggested that diversifying selection occurred after gene duplication. The fixed amino acid changes were observed in the third, fifth and sixth transmembrane domains, which has been predicted to serve as a ligand-binding pocket in a structural model. Compatibility test suggested that interlocus recombinations involving the fourth transmembrane domain occurred between the mfOR1 and mfOR2 gene pairs. The pattern of nucleotide substitutions in other OR genes agrees with the hypothesis that a limited number of amino acid residues are involved in odorant binding. Such comparative analyses of paralogous OR genes should provide bases for understanding the evolution, the structure, and the functional specificity of OR genes.

Evolutionary analysis of putative olfactory receptor genes of medaka fish, Oryzias latipes

To obtain an understanding of the origin, diversification and genomic organization of vertebrate olfactory receptor genes, we have newly cloned and characterized putative olfactory receptor genes, mfOR1, mfOR2, mfOR3 and mfOR4 from the genomic DNA of medaka fish (Oryzias latipes). The four sequences contained features commonly seen in known olfactory receptor genes and were phylogenetically most closely related to those of catfish and zebrafish. Among them, mfOR1 and mfOR2 showed the highest amino acid (aa) similarity (93%) and defined a novel olfactory receptor gene family that is most divergent among all other vertebrate olfactory receptor genes. Southern hybridization analyses suggested that mfOR1 and mfOR2 are tightly linked to each other (within 24kb), although suitable marker genes were not available to locate their linkage group. Unlike observation in catfish olfactory receptor sequences, nucleotide (nt) substitutions between the two sequences did not show any evidence of positive natural selection. mfOR3 and mfOR4, however, showed a much lower aa similarity (26%) and were both mapped to a region in the medaka linkage group XX. After including these medaka fish sequences, olfactory receptors of terrestrial and aquatic animals formed significantly different clusters in the phylogenetic tree. Although the member genes of each olfactory receptor gene subfamily are less in fish than that in mammals, fish seem to have maintained more diverse olfactory receptor gene families. Our finding of a novel olfactory receptor gene family in medaka fish may provide a step towards understanding the emergence of the olfactory receptor gene in vertebrates.

Expression Divergence of Chemosensory Genes between Drosophila sechellia and Its Sibling Species and Its Implications for Host Shift

Drosophila sechellia relies exclusively on the fruits of Morinda citrifolia, which are toxic to most insects, including its sibling species Drosophila melanogaster and Drosophila simulans. Although several odorant binding protein (Obp) genes and olfactory receptor (Or) genes have been suggested to be associated with the D. sechellia host shift, a broad view of how chemosensory genes have contributed to this shift is still lacking. We therefore studied the transcriptomes of antennae, the main organ responsible for detecting food resource and oviposition, of D. sechellia and its two sibling species. We wanted to know whether gene expression, particularly chemosensory genes, has diverged between D. sechellia and its two sibling species. Using a very stringent definition of differential gene expression, we found a higher percentage of chemosensory genes differentially expressed in the D. sechellia lineage (7.8%) than in the D. simulans lineage (5.4%); for upregulated chemosensory genes, the percentages were 8.8% in D. sechellia and 5.2% in D. simulans. Interestingly, Obp50a exhibited the highest upregulation, an approximately 100-fold increase, and Or85c—previously reported to be a larva-specific gene—showed approximately 20-fold upregulation in D. sechellia. Furthermore, Ir84a (ionotropic receptor 84a), which has been proposed to be associated with male courtship behavior, was significantly upregulated in D. sechellia. We also found expression divergence in most of the chemosensory gene families between D. sechellia and the two sibling species. Our observations suggest that the host shift of D. sechellia was associated with the enrichment of differentially expressed, particularly upregulated, chemosensory genes.

Transcriptional profiling of adult Drosophila antennae by high-throughput sequencing

BackgroundAntennae of fruit flies are the major organs responsible for detecting environmental volatiles, e.g., egg-laying substrates. An adult antenna contains many sensilla full of olfactory sensory neurons, where olfactory receptor (Or) genes are expressed. Each sensory neuron only expresses up to three receptors, making it difficult to estimate expression levels by conventional methods. In this study, we applied Illumina RNA sequencing (RNA-seq) to study the expression levels of Or and other genes in fly antennae.ResultsRNA from approximately 1,200 pairs of adult antennae from each sex of Drosophila melanogaster was used to obtain the antennal transcriptome of each sex. We detected approximately 12,000 genes expressed in antennae of either sex. The most highly expressed genes included pheromone-binding genes, transmembrane transporter genes, and sensory reception genes. Among the 61 annotated Or genes, we observed 53 and 54 genes (approximately 90%) expressed (fragments per kilobase of exon per million fragments mapped (FPKM) > 0.05) in male and female antennae, respectively; approximately 25 genes were expressed with FPKM > 15. Compared to previous studies, which extracted RNA from the whole body or head and used microarrays, antenna-specific transcriptomes obtained by RNA-seq provided more reliable estimates of gene expression levels and revealed many lowly expressed genes. Ninty-one genes, including one odorant-binding protein (Obp) gene and four Or genes, were differentially expressed between male and female antennae. These sexually biased genes were enriched on the X chromosome and showed enrichment in different gene ontology categories for male and female flies. The present and previous data together suggest that a gene family with putative immune response functions is related to pheromone detection and involved in the courtship behavior of male flies.ConclusionsTissue-specific RNA-seq is powerful for detecting lowly expressed genes. Our study provides new insight into the expression of olfactory-related genes in Drosophila antennae.

Seasonal Changes in the Long-Distance Linkage Disequilibrium in Drosophila melanogaster

Seasonal environmental changes have the potential to influence the genetic structure of species with a short generation time, such as Drosophila. We previously found the seasonal change in linkage disequilibrium (LD) between the chemoreceptor (Cr) genes in a local Japanese population (Kyoto [KY]). This could be caused by fluctuation in the population size or selection in temporally heterogeneous environments or both. Here, we analyzed the scale of LD between 51 X-linked polymorphisms (10 Cr and 41 non-Cr gene markers) in the 2 seasonal samples from the KY population and an autumn sample from 106 localities in and around Japan (Ja03au). Many of the non-Cr genes have receptor function but fewer functional connections to each other. The magnitude of LD in Ja03au did not significantly differ from that in the KY autumn sample. The lack of local differentiation was confirmed in an autumn sample from another local Japanese population. On the other hand, the magnitude of LD was significantly larger in spring than in autumn in the 2 independent KY samples. This suggests that reduction in the population size during winter increased the magnitude of LD in spring in the mainland population in Japan. Long-distance LD could be a useful measure for assessing seasonal fluctuation in effective population size.

A new test for detecting ongoing selection

Much effort has been made to search for signatures of past natural selection in DNA sequences. However, currently acting selection is rarely detected in natural populations because of its rarity, low detection power of available methods, or both. Here, we develop a new test to detect viability selection over a single generation. In this test, one specific type of chromosomes is chosen as a reference, while all other chromosomes are designated as “focal”. The test compares measures of variation between two groups of “focal” chromosomes: those found in reference/focal heterozygous individuals and those found in focal/focal homozygous individuals. In the absence of selection, we do not expect differences between these two groups as long as mating is random. On the other hand, currently acting selection can cause differences in some measures of variation. We applied this test to typing data for In(2L)t inversion polymorphism in a Drosophila melanogaster population, using “standard” (non-inverted) chromosomes as the focal class. Although the frequencies of In(2L)t and standard chromosomes did not deviate from the Hardy–Weinberg equilibrium, we found differences in allele frequency and the number of haplotypes between the two groups of standard chromosomes. This new test, in conjunction with the Hardy–Weinberg test, may shed light on how often strong selection is operating in extant populations.