Masao Watanabe - Molecular Genetics - nabe@ (Please add "ige.tohoku.ac.jp") Adjunct Professor United Graduate School of Agricultural Sciences Professor Graduate School of Life Sciences, Tohoku University Group Leader Strategic mechanism of biological systems for thermal energy - Molecular mechanism of plant reproduction under cold environment -

I. Research achievements
1. Dissection of the self/no-self recognition mechanism of self-incompatibility (SI) in Brassica species.
The self-incompatibility in higher plants is a phenomenon unique to plants that has been drawing attention since the time of C. Darwin. From the viewpoint of basic science, the self-incompatibility of cruciferous plants is a focus of attention as one of self or non-self recognition mechanisms by higher plants. The character of SI is also important for breeding practices of stable F1 hybrids. The SI of Brassica campestris is sporophytically controlled by a single locus, S, and it occurs when the S-phenotype of the pollen is the same as that of the stigma. In spite of the genetic character and their application of SI, male and female S-determinants on the S-gene, which regulates the SI recognition, had not been discovered.

By using a cruciferous plant, B. campestris, as a material plant, we started to analyze the genes encoding S-determinants at the S-locus, which regulates self/non-self recognition in SI. First, in order to determine whether SLG or SRK, which were stigma-specific S locus gene, was the female S-determinant, we constructed transgenic Brassica plants having SLG and SRK, independently. As a result, we found out that SRK was female S determinant, and that SLG acts to promote a full manifestation of the SI response. The series of our experiments for the determination of female S-determinant have been published in Nature, an international science journal.

As for the male S-determinant, we determined the whole nucleotide sequence of 76kb-genomic fragment of S locus containing SLG and SRK, and identified an anther-specific gene, SP11, having S-allelic polymorphisms. Then, in order to determine whether SP11 is a real male S-determinant or not, we constructed a transgenic plant having SP11. From the change of the pollen S-phenotype, we demonstrated that SP11 was indeed the male S-determinant, and published these results in the international journal, ProNAS. In order to examine functional and evolutionary properties of SP11, we cloned more than 20 alleles of SP11 of B. campestris, and carried out sequence analyses. The phylogenetic trees suggest possible co-evolution of the genes encoding the male and female S determinants. In addition, we found that SRK and SP11 directly interact with each other in an S haplotype-specific manner. This result was published in Nature.

We also proved that the dominance relationship among S-alleles is determined by SRK itself at the stigma side, and by the transcriptional level of SP11 at the pollen side.

Functional analysis of reproductive-organ-specific genes isolated with cDNA microarray experiments2.
Reproductive-organ-specific genes associated with the maturation of floral organs are thought to play an important role in the subsequent process of pollination and fertilization responses. Functional genome researches from these viewpoints, however, have not been carried out yet.

We produced cDNA microarray derived from the floral organs of Lotus japonicus as a model legume, and exhaustively isolated the male and female reproductive-organ-specific genes. Based on the homology of the nucleotide sequence, synchrony of expression patterns, temporal and spatial regulation of gene expression, we estimated the functions of these reproductive-organ-specific genes. These analyses revealed that many of the genes that are specifically expressed in the mature pollen grain include a group of genes that are considered to be important to the pollen tube elongation (i.e., genes associated with sugar transporting, plasma membrane re-organization, and actin re-arrangement). Also, many new tapetum cell-specific genes were isolated, suggesting the relationships to maturation of pollen surface and pollination responses.

II. Goals in the COE Program
Reproductive properties of plants are sensitive to adverse conditions, such as low temperature. While continuing to identify the genes that regulate reproduction of higher plants, we will expand the range of the study to include signal transduction mechanisms and cross talks of signal transduction between the reproductive process and low-temperature responses. In addition, influence of low-temperature-induced stimuli on reproductive properties will be analyzed on a genetic level.

III. Related website
http://news7a1.atm.iwate-u.ac.jp/~PLT-BRD/top.html

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