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National Institute for Basic Biology


Associate Professor:
Mitsuyasu Hasebe
Post Doctral Fellow:
Rumiko Kofuji1
Graduate Students:
Satomi Shindo
Yuji Hiwatashi
Satoko Kobayashi-Arakawa
Ryosuke Sano (Chiba Univ.)
Tomoaki Nishiyama (Univ. of Tokyo)
Kumi Aso (Univ. of Tokyo)
Fernando-Rivadavia Lopes (Univ. of Tokyo)
Kazuyoshi Someya (Univ. of Tokyo)
Keiko Sakakibara (Univ. of Tokyo)
Saiko Himi (Kanazawa Univ.)
Youichi Tanabe (Chiba Univ.)
Technical Staff:
Yukiko Tanikawa
Masae Umeda
Chigusa Ono

All living organisms evolved from a common ancestor more than 35 billion years ago, and accumulated mutations on their genomes caused the present biodiversity. The traces of evolutionary processes are remained in the genomes of extant organisms and we can infer (1) the phylogenetic relationships of organisms and (2) the genetic changes having caused the phenotypic evolution by comparing the genomes of different organisms. The inferred phylogenetic relationships give important insights on problems in various fields of evolutionary biology and our group is now focusing on biogeography, evolution of morphological traits and systematics in wide range of taxa. On the phenotypic evolution, we are especially interested in the morphological evolution and aim to explore genetic changes led the evolution of plant body plan. We selected Arabidopsis (angiosperm), Gnetum (gymnosperm),Ginkgo (gymnosperm), Ceratopteris (pteridophyte), and Physcomitrella (bryophyte) as models to compare the genetic cascades regulating morphogenesis especially in the reproductive organs and shoot apical meristem of land plants.

I. Molecular phylogeny of plants and lichen

(a) Phylogeny of maple trees and their disjunct distribution between North America and eastern Asia.

The maple tree genus, Acer is the largest genus in broad-leaved deciduous trees and contains about 200 species. The delimitation of the genus is clear but the intrageneric classification was controversial because of homoplasies in morphological characters. The phylogenetic relationship in Acer was inferred based on chloroplast DNA restriction site polymorphisms with 17 restriction endonucleases and previously proposed intrageneric classifications were evaluated. Based on the estimated evolutionary rate of chlotoplast DNA, divergence of eastern Asian and North American species in two different sections were estimated to have taken place in late Miocene. In consideration with previous data, multiple migrations and disjunctions are likely to have formed the eastern Asian and North American similar but disjunct distribution.

(b) Phylogeny of Coriaria and the biogeography.

Coriaria is distributed in four separate areas in the world, which is the most conspicuous disjunct distribution in flowering plants. The phylogenetic relationships of 12 Coriaria species collected from the representative disjunct areas were inferred by comparing the combined data set of rbcL (a large subunit of ribulose 1,5-bisphosphate carboxylase / oxygenase) and matK (maturase K) genes. The estimated divergence time between the Eurasian species and other species distributed in the Southern Hemisphere was estimated as 63 and 59 million years ago using rbcL and matKmolecular clocks, respectively. These results do not support the previously proposed hypotheses to explain the disjunct distribution based on the continental drift, but suggest that the distribution pattern was formed by several geographical migrations and separations in the Cenozoic.

(c) Phylogeny of lady ferns.

The lady fern group, Physematieae is one of five tribes in the Dryopteridaceae and contains about 700 species distributed mainly in temperate forests. The classification of the group is controversial and nucleotide sequences of the chloroplast gene rbcL from 42 species of the fern tribe Physematieae (Dryopteridaceae) were analyzed to provide insights into the inter and intra generic relationships and the generic circumscriptions of the group. Phylogenetic relationships of several enigmatic genera were revealed.

(d) Other phylogenetic studies

The Hydrostachiacear and the Podostemaceae have unusual morphology in angiosperms and their phylogentic relationships are controversial. During the course of field expedition to Madagascar (August to September, 1998), more than 10 species of Hydrostachis and three species of Podostemaceae were collected and the phylogenetic analyses are in progress. Molecular phylogeny of Myelochroa (lichen), Diplazium (ferns), major groups in bryophytes, and the Droseraceae (carnivorous plants) are also in progress.

II. Evolution of body plan in plants

(a) Evolution of reproductive organs.

A flower is the most complex reproductive organ in land plants and composed of sepals, petals, stamens, and gynoecium (s). Female haploid reproductive cells are covered with a sporangium (nucellus) and two integuments, and further enclosed in a gynoecium. Male haploid reproductive cells (pollens) are covered with a sporangium (pollen sack). On the other hand, gymnosperms and ferns have simpler reproductive organs than angiosperms and lack sepals and petals. Female sporangia (nucellus) of gymnosperms are covered with only one integument. Sporangia of ferns have no integuments and are naked on the abaxial side of a leaf.
The development of floral organs is mainly regulated by the members of the MADS gene family whose members are transcription factors containing the conserved MADS and K domains. MADS genes of angiosperms are divided into more than 10 groups based on the gene tree.
What kind of changes of the MADS genes caused the evolution of the complex reproductive organs in the flowering plant lineage? To address the function of the MADS genes in a plant with primitive reproductive structures lacking specialized floral organs, MADS genes were newly sought from the fern Ceratopteris richardii. A MADS gene tree, incorporating the seed plant and fern MADS genes, revealed that the known fern MADS genes form only three gene groups. The number is considerably less than the number of MADS gene groups in seed plants. The expression patterns of five genes, representing all three fern MADS gene groups, during sporophyte and gametophyte development were examined. In flowering plants, some MADS genes are expressed in specific floral organ primordia as homeotic selector genes, while other MADS genes are expressed in both reproductive and vegetative organs. Like the latter type of flowering plant MADS genes, most of the fern MADS genes are expressed in both reproductive and vegetative organs. The ubiquitously-expressed MADS genes may be more primitive than the reproductive organ-specific MADS genes. If so, it is likely that some of the MADS genes were co-opted as homeotic selector genes of specialized reproductive organs and their expression restricted to specific floral organ primordia, events that occurred after the divergence of ferns and angiosperms. To verify the hypothesis, analyses of MADS genes in gymnosperms (Gnetum and Ginkgo) with more complexed reproductive organs than ferns, but simpler ones than angiosperms, are in progress. The restriction of MADS gene expression may have been caused by the evolution of other genes that regulate the MADS genes. LEAFY gene is one of the regulators of Arabidopsis floral homeotic MADS genes. The LEAFY gene homolog of Gnetum has been cloned and the characterization is in progress. The roles of MADS genes in non-flowering plants are mysterious and characterization of MADS gene functions in the moss Physcomitrella patens will give insights for the point.

(b) Establishment of tagged mutant library of the moss Physcomitrella patens.

Mosses have the different body plan from flowering plants. Leafy shoots of mosses are similar to the ones of angiosperms, but develop in the gametophytic generation instead of the sporophytic generation as angiosperms. Organs of mosses are much simpler than flowering plants. For example, the leaves are composed of one layer of cells. Therefore, the body plan of mosses may be regulated by different genes from angiosperms. In addition to analyses of the homologs of angiosperm genes governing morphogenesis, it is necessary to screen specific genes in the moss. We established enhancer and gene trap lines and tagged mutant libraries of Physcomitrella patens to clone genes related to leafy shoot development. P. patens is known by its high rate of homologous recombination and suitable for molecular biological analyses using the gene targeting. Our libraries should be also useful for other purposes.
Fig. 1.
Phylogeny of Acer. (A) A. albopurpurascens, (B) A. laevigatum, (C) A. pseudosieboldianum. (A) and (B) have been classified in the same group because of their peculiar simple leaves in maple trees. Phylogenetic tree based on RFLPs of chloroplast genome revealed that species with simple leaves (A and B) are polyphyletic and (B) is more closely related to the species with palmate leaves (C).
Fig. 2.
Patterns of CMADS1 RNA expression in sporophyte tissues as detected by in situ hybridization. MADS genes of Ceratopteris richardii are expressed in both reproductive (C) and vegetative (D, E, and F) tissues. (A) C. richardii sporophyte. A whole plant with reproductive (r) and vegetative leaves. (B) Naked sporangia (sp) on the abaxial surface of a reproductive leaf. (C) Longitudinal section of a sporophyll showing sporangia (sp). (D) Longitudinal section of the growing tip of a root (ro). (E) Transverse section of a petiole showing vascular bundles (v). (F) A longitudinal (l) and a transverse (t) section of two young vegetative leaves.

Publication List:
Hasebe, M., Ando, T. and Iwatsuki, K. (1998) Intrageneric relationships of maple trees based on the chloroplast DNA restriction fragment length polymorphisms. J. Plant Res. 111: 441-451.
Hasebe, M., Wen, C.-K., Kato, M. and Banks, J.A. (1998) Characterization of MADS homeotic genes in the fern Ceratopteris richardii. Proc. Natl. Acad. Sci. USA 95: 6222-6227.
Wolf, P.G., Pryer, K.M., Smith, A.R. and Hasebe, M. (1998) Phylogenetic studies of extant Pteridophytes. In D. Soltis et al. eds, Molecular Systematics of Plants (2nd), Chapmann and Hall, New York. Pp. 541-556