Nucleotide sequences from four chloroplast genes, the matK, chlL, intergenic spacer (IGS) region between trnL and trnF, and an intron of trnL, were determined from all species of Taxodiaceae and five species of Cupressaceae sensu stricto (s.s.). Phylogenetic trees were constructed using the maximum parsimony and the neighbor-joining methods with Cunninghamia as an outgroup. These analyses provided greater resolution of relationships among genera and higher bootstrap supports for clades compared to previous analyses. Results indicate that Taiwania diverged first, and then Athrotaxis diverged from the remaining genera. Metasequoia, Sequoia, and Sequoiadendron form a clade. Taxodium and Glyptostrobus form a clade, which is the sister to Cryptomeria. Cupressaceae s.s. are derived from within Taxodiaceae, being the most closely related to the Cryptomeria/Taxodium/Glyptostrobus clade. These relationships are consistent with previous morphological groupings and the analyses of molecular data. In addition, we found acceleration of evolutionary rates in Cupressaceae s.s. Possible causes for the acceleration are discussed.

Sleep behaviors are observed even in nematodes and arthropods, yet little is known about how sleep-regulatory mechanisms have emerged during evolution. Here, we report a sleep-like state in the cnidarian Hydra vulgaris with a primitive nervous organization. Hydra sleep was shaped by homeostasis and necessary for cell proliferation, but it lacked free-running circadian rhythms. Instead, we detected 4-hour rhythms that might be generated by ultradian oscillators underlying Hydra sleep. Microarray analysis in sleep-deprived Hydra revealed sleep-dependent expression of 212 genes, including cGMP-dependent protein kinase 1 (PRKG1) and ornithine aminotransferase. Sleep-promoting effects of melatonin, GABA, and PRKG1 were conserved in Hydra. However, arousing dopamine unexpectedly induced Hydra sleep. Opposing effects of ornithine metabolism on sleep were also evident between Hydra and Drosophila, suggesting the evolutionary switch of their sleep-regulatory functions. Thus, sleep-relevant physiology and sleep-regulatory components may have already been acquired at molecular levels in a brain-less metazoan phylum and reprogrammed accordingly.

DNA-based transposable elements can be used as tools for gene engineering and gene therapy. A great advantage over RNA-mediated elements and retroviruses is the simplicity and safety of usage. The Tol1 element of the medaka fish Oryzias latipes has structural features of DNA-based elements. Although its excision has already been demonstrated, de novo insertion has not been observed, and a transposase has not been hitherto identified. We first cloned, through in silico search alignments and genomic library screenings, a 4.4-kb Tol1 copy carrying open reading frames and then identified, by mRNA analysis, a 2.9-kb transcript coding for 851 amino acids. The protein product of this transcript catalyzed transposition of a nonautonomous Tol1 copy in human and mouse culture cells. This identification of a fully functional Tol1 transposase could lead to the development of new tools for basic and translational molecular biology applications in mammals.

Sympatric speciation has been demonstrated in few empirical case studies, despite intense searches, because of difficulties in testing the criteria for this mode of speciation. Here, we report a possible case of sympatric speciation in ricefishes of the genus Oryzias on Sulawesi, an island of Wallacea. Three species of Oryzias are known to be endemic to Lake Poso, an ancient tectonic lake in central Sulawesi. Phylogenetic analyses using RAD‐seq‐derived single nucleotide polymorphisms (SNPs) revealed that these species are monophyletic. We also found that the three species are morphologically distinguishable and clearly separated by population‐structure analyses based on the SNPs, suggesting that they are reproductively isolated from each other. A mitochondrial DNA chronogram suggested that their speciation events occurred after formation of the tectonic lake, and existence of a historical allopatric phase was not supported by coalescent‐based demographic inference. Demographic inference also suggested introgressive hybridization from an outgroup population. However, differential admixture among the sympatric species was not supported by any statistical tests. These results all concur with criteria necessary to demonstrate sympatric speciation. Ricefishes in this Wallacean lake provide a promising new model system for the study of sympatric speciation.

We surveyed the molecular evolutionary characteristics of 11 nuclear genes from 10 conifer trees belonging to the Taxodioideae, the Cupressoideae, and the Sequoioideae. Comparisons of substitution rates among the lineages indicated that the synonymous substitution rates of the Cupressoideae lineage were higher than those of the Taxodioideae. This result parallels the pattern previously found in plastid genes. Likelihood-ratio tests showed that the nonsynonymous-synonymous rate ratio did not change significantly among lineages. In addition, after adjustments for lineage effects, the dispersion indices of synonymous and nonsynonymous substitutions were considerably reduced, and the latter was close to 1. These results indicated that the acceleration of evolutionary rates in the Cupressoideae lineage occurred in both the nuclear and plastid genomes, and that generally, this lineage effect affected synonymous and nonsynonymous substitutions similarly. We also investigated the relationship of synonymous substitution rates with the nonsynonymous substitution rate, base composition, and codon bias in each lineage. Synonymous substitution rates were positively correlated with nonsynonymous substitution rates and GC content at third codon positions, but synonymous substitution rates were not correlated with codon bias. Finally, we tested the possibility of positive selection at the protein level, using maximum likelihood models, assuming heterogeneous nonsynonymous-synonymous rate ratios among codon (amino acid) sites. Although we did not detect strong evidence of positively selected codon sites, the analysis suggested that significant variation in nonsynonymous-synonymous rate ratio exists among the sites. The most likely sites for action of positive selection were found in the ferredoxin gene, which is an important component of the apparatus for photosynthesis.

The obligate mutualism of figs and fig-pollinating wasps has been one of the classic models used for testing theories of co-evolution and cospeciation due to the high species-specificity of these relationships. To investigate the species-specificity between figs and fig pollinators and to further understand the speciation process in obligate mutualisms, we examined the genetic differentiation and phylogenetic relationships of four closely related fig-pollinating wasp species (Blastophaga nipponica, Blastophaga taiwanensis, Blastophaga tannoensis and Blastophaga yeni) in Japan and Taiwan using genome-wide sequence data, including mitochondrial DNA sequences. In addition, population structure was analysed for the fig wasps and their host species using microsatellite data. The results suggest that the three Taiwanese fig wasp species are a single panmictic population that pollinates three dioecious fig species, which are sympatrically distributed, have large differences in morphology and ecology and are also genetically differentiated. Our results illustrate the first case of pollinator sharing by host shift in the subgenus Ficus. On the other hand, there are strict genetic codivergences between allopatric populations of the two host-pollinator pairs. The possible processes that produce these pollinator-sharing events are discussed based on the level and pattern of genetic differentiation in these figs and fig wasps.

The light-independent protochlorophyllide oxidoreductase (DPOR) plays a key role in the ability of nonflowering plants and algae to synthesize chlorophyll in darkness. This enzyme consists of three subunits encoded by the chlB, chlL, and chlN genes in the plastid genome. Previously, we found a high nonsynonymous substitution rate (dN) of the chlL gene in the lineage of Thuja standishii, a conifer belonging to the Cupressaceae. Here we revealed that the acceleration of dN in the chlL occurred as well in other species of Thuja, Thuja occidentalis and Thuja plicata. In addition, dark-grown seedlings of T. occidentalis were found to exhibit a pale yellowish color, and their chlorophyll concentration was much lower than that of other species of Cupressaceae. The results suggested that the species of Thuja have lost the ability to synthesize chlorophyll in darkness, and the functional constraint on the DPOR would thus be expected to be relaxed in this genus. Therefore, we expected to find that the evolutionary rates of all subunits of DPOR would in this case be accelerated. Sequence analyses of the chlN and chlB (encoding the other subunits of DPOR) in 18 species of Cupressaceae revealed that the dN of the chlN gene was accelerated in Thuja as was the dN of the chlL gene, but the dN of the chlB gene did not appear to differ significantly among the species of Cupressaceae. Sequencing of reverse transcription-polymerase chain reaction (RT-PCR) products of these genes showed that RNA editing was rare and unlikely to have contributed to the acceleration. Moreover, the RT-PCR analysis indicated that all chl genes were still transcriptionally active in T. occidentalis. Based on these results, it appears that species of Thuja still bear the DPOR protein, although the enzyme has lost its activity because of nonsynonymous mutations of some of the chl genes. The lack of acceleration of the dN of the chlB gene might be accounted for by various unknown functions of its gene product.

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