Relationship between reduction in rice (Nipponbare) leaf blade size under elevated CO2 and miR396-GRF module.

Elevated CO2 (eCO2; 1000 ppm) influences developing rice leaf formation, reducing leaf blade length and width as compared to rice grown under ambient CO2 (aCO2; 400 ppm). Since micro RNAs (miRNAs) are known to play multiple roles in plant development, we hypothesized that miRNAs might be involved in modulating leaf size under eCO2 conditions. To identify miRNAs responding to eCO2, we profiled miRNA levels in developing rice leaves (P4; plastochron number of the fourth-youngest leaf) under eCO2 using small RNA-seq. We detected 18 mature miRNA sequences for which expression levels varied more than two-fold between the eCO2 and aCO2 conditions. Among them, only miR396e and miR396f significantly differed between the two conditions. Additionally, the expression of growth-regulating factors (GRFs), potential target mRNA of miR396s, were repressed under the eCO2 condition. We used an antisense oligonucleotide approach to confirm that single-strand DNA corresponding to the miR396e sequence effectively downregulated GRF expression in developing leaves, reducing the leaf blade length, such as for rice grown under eCO2. These results suggest that the miR396-GRF module is crucially relevant to controlling rice leaf blade length in eCO2 environments.

Biochemical analysis of a new sugary-type rice mutant, Hemisugary1, carrying a novel allele of the sugary-1 gene.

MAIN CONCLUSION: A novel allele of the sugary-1 rice mutant was isolated. The single amino acid change led to isoamylase activity reduction and accumulation of high-molecular-weight phytoglycogen in seeds. A new sugary rice variety with an improved seed appearance has been isolated and designated Hemisugary1. This mutant, which was derived from Japonica-type cultivar Tsugaruroman treated with sodium azide, has about half the isoamylase activity of seeds in the original Tsugaruroman. The mutant also accumulates significant phytoglycogen, albeit approximately 40% of the total phytoglycogen in the existing sugary cultivar Ayunohikari which is defective in its most isoamylase activity. The site of mutation was identified using a re-sequence of the whole genome and a cleaved amplified polymorphic sequence (CAPS) marker. The hemisugary phenotypes of the F2 progeny were entirely consistent with the results of genotyping using the CAPS marker. Segregation analysis of the F2 population showed that the hemisugary phenotype was controlled by a single recessive gene, which was produced by a G → A single nucleotide polymorphism in the sugary-1 gene, resulting in a missense mutation from glycine to aspartic acid at amino acid position 333. Zymogram showed that this amino acid replacement resulted in a decrease in isoamylase activity with a concomitant reduction in the formation of isoamylase complexes. Phytoglycogen molecules from Hemisugary1 seeds were 3.5 times larger and contained more short glucan chains than did Ayunohikari seeds. Our data provide new insights into the relationship between isoamylase structure and phytoglycogen formation.

Measurement of Water Distribution on Micro-structured Surface Buried in Water as a Model of Super Water Repellent Surface by Stimulated Raman Scattering Interferometer.

Water repellency of surfaces is realized by the roughness of the surfaces and the coating compounds on the surfaces. It is known that the water repellent surfaces are due to invasion of water into the trench structures on the surface, or due to air remaining in the trench structures. However, for more effective designing and fabrication of water-repellent surfaces, it is essential to measure the spatial distribution of water around the microstructures. In the present study, the SRS interferometer developed by the authors is applied to distinguish whether water or air is present in the trenches on the water-substrate interface buried by water. By obtaining the SRS interference signal generated from water while scanning the sample position, the phase shift of the SRS interference signal due to the microstructure on the interface was observed. From the quantitative analysis of the phase difference, it was revealed that the trench was filled with water. It is concluded that SRS interferometry is an effective method to observe experimentally the interface condition with chemical contrast along microstructures.