論文 - 永野　惇

A first genetic map in the genus Streptocarpus generated with RAD sequencing based SNP markers

Chen Y., Nishii K., Barber S., Hackett C., Kidner C., Gharbi K., Nagano A., Iwamoto A., Möller M.

South African Journal of Botany （South African Journal of Botany）  117   158 - 168 2018年07月

ISSN  02546299

　概要を見る

Streptocarpus shows a great diversity of shoot and flower morphology, but lacks resources for genetic studies. RAD sequencing with NGS technologies provides large amounts of genotyping data quickly and cost effectively and is very useful for genetic map construction. In this study, three approaches for constructing RAD loci were combined to build the first genetic map for Streptocarpus (Gesneriaceae). A backcross population between Streptocarpus grandis and S. rexii was used as the mapping population. The parents are both diploid (2n = 32) and largely autogamous. Streptocarpus grandis is unifoliate, retaining the enlarged macrocotyledon as sole foliar organ which is monocarpic, and S. rexii is rosulate and perennial. 233 backcross plants were genotyped, and the data from 200 or 150 individuals used to generate the maps. The RAD reads were assembled using a de novo approach, aligned on the reference draft genome sequences of S. rexii using BWA or Stampy, and finally all markers were analyzed combined. In the latter, 599 mappable markers were generated which were assigned to 16 linkage groups, with an average inter-marker distance of 2.6 cM. This work provides a valuable genetic resource for further studies in the large and morphologically diverse genus Streptocarpus.

• Access to Document (DOI)

Functional divergence of duplicate genes several million years after gene duplication in Arabidopsis

Hanada K., Tezuka A., Nozawa M., Suzuki Y., Sugano S., Nagano A.J., Ito M., Morinaga S.I.

DNA Research （DNA Research）  25 （ 3 ） 327 - 339 2018年06月

ISSN  13402838

　概要を見る

Lineage-specific duplicated genes likely contribute to the phenotypic divergence in closely related species. However, neither the frequency of duplication events nor the degree of selection pressures immediately after gene duplication is clear in the speciation process. Here, using Illumina DNA-sequencing reads from Arabidopsis halleri, which has multiple closely related species with high-quality genome assemblies (A. thaliana and A. lyrata), we succeeded in generating orthologous gene groups in Brassicaceae. The duplication frequency of retained genes in the Arabidopsis lineage was 10 times higher than the duplication frequency inferred by comparative genomics of Arabidopsis, poplar, rice and moss (Physcomitrella patens). The difference of duplication frequencies can be explained by a rapid decay of anciently duplicated genes. To examine the degree of selection pressure on genes duplicated in either the A. halleri-lyrata or the A. halleri lineage, we examined positive and purifying selection in the A. halleri-lyrata and A. halleri lineages throughout the ratios of nonsynonymous to synonymous substitution rates (KA/ KS). Duplicate genes tended to have a higher proportion of positive selection compared with non-duplicated genes. Interestingly, we found that functional divergence of duplicated genes was accelerated several million years after gene duplication compared with immediately after gene duplication.

• Access to Document (DOI)

Auxin contributes to the intraorgan regulation of gene expression in response to shade

Kim S., Mochizuki N., Deguchi A., Nagano A., Suzuki T., Nagatani A.

Plant Physiology （Plant Physiology）  177 （ 2 ） 847 - 862 2018年06月

ISSN  00320889

　概要を見る

Plants sense and respond to light via multiple photoreceptors including phytochrome. The decreased ratio of red to far-red light that occurs under a canopy triggers shade-avoidance responses, which allow plants to compete with neighboring plants. The leaf acts as a photoperceptive organ in this response. In this study, we investigated how the shade stimulus is spatially processed within the cotyledon. We performed transcriptome analysis on microtissue samples collected from vascular and nonvascular regions of Arabidopsis (Arabidopsis thaliana) cotyledons. In addition, we mechanically isolated and analyzed the vascular tissue. More genes were up-regulated by the shade stimulus in vascular tissues than in mesophyll and epidermal tissues. The genes up-regulated in the vasculature were functionally divergent and included many auxin-responsive genes, suggesting that various physiological/developmental processes might be controlled by shade stimulus in the vasculature. We then investigated the spatial regulation of these genes in the vascular tissues. A small vascular region within a cotyledon was irradiated with far-red light, and the response was compared with that when the whole seedling was irradiated with far-red light. Most of the auxin-responsive genes were not fully induced by the local irradiation, suggesting that perception of the shade stimulus requires that a wider area be exposed to far-red light or that a certain position in the mesophyll and epidermis of the cotyledon be irradiated. This result was consistent with a previous report that auxin synthesis genes are up-regulated in the periphery of the cotyledon. Hence, auxin acts as an important intraorgan signaling factor that controls the vascular shade response within the cotyledon.

• Access to Document (DOI)

Phylogeographic testing of alternative histories of single-origin versus parallel evolution of early flowering serpentine populations of Picris hieracioides L. (Asteraceae) in Japan

Sakaguchi S., Horie K., Kimura T., Nagano A., Isagi Y., Ito M.

Ecological Research （Ecological Research）  33 （ 3 ） 537 - 547 2018年05月

ISSN  09123814

　概要を見る

Serpentine, a chemically extreme substrate with a discontinuous distribution, supports a high proportion of endemic plant species. The evolutionary pathway of edaphic endemics in isolated areas may be of parallel origin (convergence), but this can be difficult to detect due to similar sets of phenotypic traits repeatedly selected for by the extreme conditions of serpentine environments. Testing the alternative histories of single origin vs. parallel evolution in serpentine plant populations provides insight into the ability of plants to adapt to extreme serpentine environments and to disperse between isolated serpentine areas. In this study, we focused on the herb Picris hieracioides L. (Asteraceae), which includes early-flowering serpentine populations surrounded by later flowering, non-serpentine populations on Hokkaido Island, Japan. We performed population genetic analysis based on genome-wide SNPs markers and quantified temporal overlap of flowering times under natural conditions. Phylogeographic analysis confirmed the occurrence of two genetic groups, corresponding to the serpentine and non-serpentine races. Although the two races were found to occur in close parapatry over multiple paired soil sites, little evidence for hybridization was detected. The flowering times of the two races showed significant divergence, with flowering peaks separated by more than 1 month. Overall, this study supported the single origin of serpentine Picris populations, and showed the role of divergent flowering time as a strong prezygotic barrier to maintain the edaphic races over short spatial distances.

• Access to Document (DOI)

Does genomic variation in a foundation species predict arthropod community structure in a riparian forest?

Kagiya S., Yasugi M., Kudoh H., Nagano A.J., Utsumi S.

Molecular Ecology （Molecular Ecology）  27 （ 5 ） 1284 - 1295 2018年03月

ISSN  09621083

　概要を見る

Understanding how genetic variation within a foundation species determines the structure of associated communities and ecosystem processes has been an emerging frontier in ecology. Previous studies in common gardens identified close links between intraspecific variation and multispecies community structure, and these findings are now being evaluated directly in the complex natural ecosystem. In this study, we examined to what extent genomic variation in a foundation tree species explains the structure of associated arthropod communities in the field, comparing with spatial, temporal and environmental factors. In a continuous mixed forest, arthropods were surveyed on 85 mature alders (Alnus hirsuta) in 2 years. Moreover, we estimated Nei's genetic distance among the alders based on 1,077 single nucleotide polymorphisms obtained from restricted-site-associated DNA sequencing of the alders’ genome. In both years, we detected significant correlations between genetic distance and dissimilarity of arthropod communities. A generalized dissimilarity modelling indicated that the genetic distance of alder populations was the most important predictor to explain the variance of arthropod communities. Among arthropod functional groups, carnivores were consistently correlated with genetic distance of the foundation species in both years. Furthermore, the extent of year-to-year changes in arthropod communities was more similar between more genetically closed alder populations. This study demonstrates that the genetic similarity rule would be primarily prominent in community assembly of plant-associated arthropods under temporally and spatially variable environments in the field.

• Access to Document (DOI)

Phylogeographic analysis of the East Asian goldenrod (Solidago virgaurea complex, Asteraceae) reveals hidden ecological diversification with recurrent formation of ecotypes

Sakaguchi S., Kimura T., Kyan R., Maki M., Nishino T., Ishikawa N., Nagano A.J., Honjo M.N., Yasugi M., Kudoh H., Li P., Choi H.J., Chernyagina O.A., Ito M.

Annals of Botany （Annals of Botany）  121 （ 3 ） 489 - 500 2018年02月

ISSN  03057364

　概要を見る

• Background and Aims: The processes and mechanisms underlying lineage diversification are major topics in evolutionary biology. Eurasian goldenrod species of the Solidago virgaurea complex show remarkable morphological and ecological diversity in the Japanese Archipelago, with ecotypic taxa well adapted to specific environments (climate, edaphic conditions and disturbance regimes). The species complex is a suitable model to investigate the evolutionary processes of actively speciating plant groups, due to its ability to evolve in relation to environmental adaptation and its historical population dynamics. • Methods: Two chloroplast markers, 18 nuclear microsatellite markers and ddRAD-sequencing were used to infer population genetic demography of S. virgaurea complex with its related species/genera. • Key Results: Our analysis showed that populations in Japan form an evolutionary unit, which was genetically diverged from adjacent continental populations. The phylogenetic structure within the archipelago strongly corresponds to the geography, but interestingly there is no concordance between genetic structure and ecotypic boundaries; neighbouring populations of distinct ecotypes share a genetic background. • Conclusions: We propose that the traits specific to the ecotypic entities are maintained by natural selection or are very recently generated and have little effect on the genomes, making genome-wide genetic markers unsuitable for detecting ecotypic differentiation. Furthermore, some sporadically distributed taxa (found as rheophytes and alpine plants) were repeatedly generated from a more widespread taxon in geographically distant areas by means of selection. Overall, this study showed that the goldenrod complex has a high ability to evolve, enabling rapid ecological diversification over a recent timeframe.

• Access to Document (DOI)

Phylogenetic relationships of Aurantioideae (Rutaceae) based on RAD-Seq

Nagano Y., Mimura T., Kotoda N., Matsumoto R., Nagano A.J., Honjo M.N., Kudoh H., Yamamoto M.

Tree Genetics and Genomes （Tree Genetics and Genomes）  14 （ 1 ）  2018年02月

ISSN  16142942

　概要を見る

The economically and nutritionally important genus Citrus belongs to the subfamily Aurantioideae in the family Rutaceae. Here, we analyzed the phylogenetic relationships of the subfamily Aurantioideae based on RAD-Seq. The RAD-Seq data produced phylogenetic trees with high support values, clear discriminations based on branch length, and elucidations of early branching events. Our genetic classification corresponded well with the classical morphological classification system and supported the subdivision of Citreae, one of two tribes of the Aurantioideae, into three subtribes—Triphasiinae, Citrinae, and Balsamocitrinae. Additionally, it was largely consistent with the subdivision of Clauseneae, the other tribe of the Aurantioideae, into three subtribes—Micromelinae, Clauseninae, and Merrillinae; the exception was Murraya paniculata. With the exception of members of primitive citrus fruit trees, namely, Severinia buxifolia and Hesperethusa crenulata, lower-level morphological groupings under subtribes based on genetic and morphological classifications corresponded well. The phylogenetic relationship between Asian “true citrus fruit trees” (genera Citrus, Poncirus, and Fortunella) and Australian/New Guinean citrus fruit trees (genera Microcitrus, Eremocitrus, and Clymenia) was inconsistent between present classification based mainly on the nuclear genome and the previous classification based on the chloroplast genome. This inconsistency may be explained by chloroplast capture. Our findings provide a valuable insight into the genetic relationships of the subfamily Aurantioideae in the family Rutaceae.

• Access to Document (DOI)

Estimation of the circadian phase by oscillatory analysis of the transcriptome in plants

Takeoka M., Higashi T., Honjo M., Tezuka A., Nagano A., Tanigaki Y., Fukuda H.

Environmental Control in Biology （Environmental Control in Biology）  56 （ 2 ） 67 - 72 2018年

ISSN  1880554X

　概要を見る

The phase of the circadian clock (body time) is related to various aspects of physiology and metabolism in plants. Therefore, it is important to evaluate body time to control plant quality and growth. The molecular timetable method, a statistical analysis of the transcriptome, is useful for plant body time estimation. However, it is necessary to verify some assumptions in this method for high resolution estimation in plants under various conditions. To investigate whether the assumptions are valid, we analyzed the period length and expression pattern of genes. We used Arabidopsis thaliana as a model plant, and both lettuce and tomato as model crops. A. thaliana and lettuce were cultivated under conditions of constant light in a closed plant factory, and tomato was cultivated in a sunlight-type plant factory. We found that estimation of the period length of the plant circadian rhythm was possible and that the gene expression pattern correlates well with test-fit cosine curves. Thus, we successfully demonstrated high-resolution estimation of body time in plants using a molecular timetable method.

• Access to Document (DOI)

Growth and environmental change-independent genes associated with clock gene TOC1 in green perilla

Tanigaki Y., Higashi T., Nagano A., Honjo M., Fukuda H.

Environmental Control in Biology （Environmental Control in Biology）  56 （ 4 ） 137 - 142 2018年

ISSN  1880554X

　概要を見る

Secondary metabolism is regulated by circadian rhythm based on the clock genes expression. Transcriptome analysis is an important tool to understand these mechanisms. However, sufficient genetic information for this analysis is lacking for many crops. In order to analyze the influence of independent clock genes from growth and environmental stimulation, we investigated the gene expression profiles in the roots and leaves of green perilla (Perilla frutescens), which is an important crop used as a herbal medicine. Green perilla was grown under light emitting diode and fluorescent light, after 7, 14, and 35 d. We also compared the coding DNA sequences with those in the other plants. From the transcriptomic data, green perilla was observed to have 74 contigs that did not depend on growth and cultivation condition. Some of these 74 genes might be related to the clock genes through their interactions with other genes. Some 74 genes had TATA box-like motif and a GA motif of the genes that are regulatory targets of timing of CAB expression 1 (TOC1). Thus, 74 genes that did not depend on cultivation condition and growth were considered to be under the control of TOC 1, suggesting the importance of TOC1 in circadian rhythms.

• Access to Document (DOI)

Selection of transcripts affecting initial growth rate of rice backcrossed inbred lines using RNA sequencing data

Fukuda A., Hirose T., Aoki N., Kondo S., Yonekura M., Kataoka T., Ohto C., Nagano A.

Frontiers in Plant Science （Frontiers in Plant Science）  871 2018年

　概要を見る

Seedling growth is an important factor for direct seeding of rice. However, the genetic and transcriptomic factors involved in this process are largely unknown. In this study, transcripts affecting shoot weight were identified in rice (Oryza sativa L.) using RNA sequencing (RNA-Seq) data from 20 backcrossed inbred lines (BILs) and their parental cultivars. The selection frequency of the genes for the regression model was determined using repeated analysis of random subsets of the transcriptome. The qLTG3-1gene, controlling low-temperature germinability, and short grain 1 gene (SG1), known to decrease organ elongation, showed high frequency. The quantitative trait loci (QTLs) analysis performed for BILs revealed that qLTG3-1 was included in the QTLs for shoot weight but SG1 was not. No nucleotide polymorphisms were found in the coding region of SG1 in either of the parental cultivars. Quantitative real-time PCR showed that SG1 expression was negatively correlated with shoot weight for all 104 BILs analyzed in this study. Expression QTL (eQTLs) analysis showed an eQTL for SG1 expression located in the same region as the QTL for shoot weight. However, no eQTLs were detected on the same chromosome as SG1, suggesting that nucleotide polymorphisms around the gene do not affect its expression in analyzed growth stage. Overall, these results indicate that RNA-Seq is a useful tool for identifying transcripts that can be related to seedling growth rate.

• Access to Document (DOI)