Jinmi Yoon

OsPUB15, an E3 ubiquitin ligase, functions to reduce cellular oxidative stress during seedling establishment

The plant U-box (PUB) protein functions as an E3 ligase to poly-ubiquitinate a target protein for its degradation or post-translational modification. Here, we report functional roles for OsPUB15, which encodes a cytosolic U-box protein in the class-II PUB family. Self-ubiquitination assays showed that bacterially expressed MBP-OsPUB15 protein has E3 ubiquitin ligase activity. A T-DNA insertional mutation in OsPUB15 caused severe growth retardation and a seedling-lethal phenotype. Mutant seeds did not produce primary roots, and their shoot development was significantly delayed. Transgenic plants expressing the OsPUB15 antisense transcript phenocopied these mutant characters. The abnormal phenotypes were partially rescued by two antioxidants, catechin and ascorbic acid. Germinating seeds in the dark also recovered the rootless defect. Levels of H2O2 and oxidized proteins were higher in the knock-out mutant compared with the wild type. OsPUB15 transcript levels were increased upon H2O2, salt and drought stresses; plants overexpressing the gene grew better than the wild type under high salinity. These results indicate that PUB15 is a regulator that reduces reactive oxygen species (ROS) stress and cell death.

Mutation in Wilted Dwarf and Lethal 1 (WDL1) causes abnormal cuticle formation and rapid water loss in rice

Epidermal cell layers play important roles in plant defenses against various environmental stresses. Here we report the identification of a cuticle membrane mutant, wilted dwarf and lethal 1 (wdl1), from a rice T-DNA insertional population. The muant is dwarf and die at seedling stage due to increased rates of water loss. Stomatal cells and pavement cells are smaller in the mutant, suggesting that WDL1 affects epidermal cell differentiation. T-DNA was inserted into a gene that encodes a protein belonging to the SGNH subfamily, within the GDSL lipase superfamily. The WDL1–sGFP signal coincided with the RFP signal driven by AtBIP–mRFP, indicating that WDL1 is an ER protein. SEM analyses showed that their leaves have a disorganized crystal wax layer. Cross-sectioning reveals loose packing of the cuticle and irregular thickness of cell wall. Detailed analyses of the epicuticular wax showed no significant changes either in the total amount and amounts of each monomer or in the levels of lipid polymers, including cutin and other covalently bound lipids, attached to the cell wall. We propose that WDL1 is involved in cutin organization, affecting depolymerizable components.

Roles of lignin biosynthesis and regulatory genes in plant development.

Abstract Lignin is an important factor affecting agricultural traits, biofuel production, and the pulping industry. Most lignin biosynthesis genes and their regulatory genes are expressed mainly in the vascular bundles of stems and leaves, preferentially in tissues undergoing lignification. Other genes are poorly expressed during normal stages of development, but are strongly induced by abiotic or biotic stresses. Some are expressed in non‐lignifying tissues such as the shoot apical meristem. Alterations in lignin levels affect plant development. Suppression of lignin biosynthesis genes causes abnormal phenotypes such as collapsed xylem, bending stems, and growth retardation. The loss of expression by genes that function early in the lignin biosynthesis pathway results in more severe developmental phenotypes when compared with plants that have mutations in later genes. Defective lignin deposition is also associated with phenotypes of seed shattering or brittle culm. MYB and NAC transcriptional factors function as switches, and some homeobox proteins negatively control lignin biosynthesis genes. Ectopic deposition caused by overexpression of lignin biosynthesis genes or master switch genes induces curly leaf formation and dwarfism.

The control of flowering time by environmental factors

The timing of flowering is determined by endogenous genetic components as well as various environmental factors, such as day length, temperature, and stress. The genetic elements and molecular mechanisms that rule this process have been examined in the long-day-flowering plant Arabidopsis thaliana and short-day-flowering rice (Oryza sativa). However, reviews of research on the role of those factors are limited. Here, we focused on how flowering time is influenced by nutrients, ambient temperature, drought, salinity, exogenously applied hormones and chemicals, and pathogenic microbes. In response to such stresses or stimuli, plants either begin flowering to produce seeds for the next generation or else delay flowering by slowing their metabolism. These responses vary depending upon the dose of the stimulus, the plant developmental stage, or even the cultivar that is used. Our review provides insight into how crops might be managed to increase productivity under various environmental challenges.

KNOX Protein OSH15 Induces Grain Shattering by Repressing Lignin Biosynthesis Genes

Major domestication factors for grain shattering in rice, qSH1 and SH5, interact with binding partner OSH15 KNOX protein to control abscission zone development and repress lignin biosynthesis. Seed shattering is an agronomically important trait. Two major domestication factors are responsible for this: qSH1 and SH5. Whereas qSH1 functions in cell differentiation in the abscission zone (AZ), a major role of SH5 is the repression of lignin deposition. We have determined that a KNOX protein, OSH15, also controls seed shattering. Knockdown mutations of OSH15 showed reduced seed-shattering phenotypes. Coimmunoprecipitation experiments revealed that OSH15 interacts with SH5 and qSH1, two proteins in the BELL homeobox family. In transgenic plants carrying the OSH15 promoter-GUS reporter construct, the reporter gene was preferentially expressed in the AZ during young spikelet development. The RNA in situ hybridization experiment also showed that OSH15 messenger RNAs were abundant in the AZ during spikelet development. Analyses of osh15 SH5-D double mutants showed that SH5 could not increase the degree of seed shattering when OSH15 was absent, indicating that SH5 functions together with OSH15. In addition to the seed-shattering phenotype, osh15 mutants displayed dwarfism and accumulated a higher amount of lignin in internodes due to increased expression of the genes involved in lignin biosynthesis. Knockout mutations of CAD2, which encodes an enzyme for the last step in the monolignol biosynthesis pathway, caused an easy seed-shattering phenotype by reducing lignin deposition in the AZ. This indicated that the lignin level is an important determinant of seed shattering in rice (Oryza sativa). Chromatin immunoprecipitation assays demonstrated that both OSH15 and SH5 interact directly with CAD2 chromatin. We conclude that OSH15 and SH5 form a dimer that enhances seed shattering by directly inhibiting lignin biosynthesis genes.

Identification of root-preferential transcription factors in rice by analyzing GUS expression patterns of T-DNA tagging lines

T-DNA tagging lines are useful for analyzing the functions of genes and regulatory elements. We have previously generated approximately 100,000 insertional mutants in japonica rice (Oryza sativa), using T-DNA vectors carrying the promoter-less GUS reporter gene. In this study, we conducted GUS assays of seedlings from 430 lines in which TDNA was inserted into transcription factor genes. Among the 75 lines that showed GUS signals, nine displayed an endospermpreferential expression pattern; two lines demonstrated GUS signals in both endosperm and roots; 21 lines had GUS expression mainly in leaves; 19 lines showed GUS signal in both leaves and roots; and 24 lines expressed GUS predominantly in the roots. Co-segregation analyses of 49 homozygous lines indicated that the GUS expression patterns observed from 38 lines were due to the T-DNA insertion. We also identified fusion transcripts between tagged genes and the GUS reporter in six lines. Quantitative RT-PCR confirmed that the GUS expression patterns of those tagged lines indeed represent organ- and tissue-preferential expression of the tagged genes. The GUS-tagged transcription factor lines identified here will be useful for functional analysis of these candidates.

Chromatin interacting factor OsVIL2 increases biomass and rice grain yield

Summary Grain number is an important agronomic trait. We investigated the roles of chromatin interacting factor Oryza sativa VIN3‐LIKE 2 (OsVIL2), which controls plant biomass and yield in rice. Mutations in OsVIL2 led to shorter plants and fewer grains whereas its overexpression (OX) enhanced biomass production and grain numbers when compared with the wild type. RNA‐sequencing analyses revealed that 1958 genes were up‐regulated and 2096 genes were down‐regulated in the region of active division within the first internodes of OX plants. Chromatin immunoprecipitation analysis showed that, among the downregulated genes, OsVIL2 was directly associated with chromatins in the promoter region of CYTOKININ OXIDASE/DEHYDROGENASE2 (OsCKX2), a gene responsible for cytokinin degradation. Likewise, active cytokinin levels were increased in the OX plants. We conclude that OsVIL2 improves the production of biomass and grain by suppressing OsCKX2 chromatin.

Roles of Sugars in Controlling Flowering Time

Flowering time is influenced by environmental factors such as photosynthesis, temperature, nutrition, and water. The main products of photosynthesis are sugars that are mobilized to sink tissues to support plant growth and differentiation. They also function as signals to control various types of metabolism and developmental processes. One of the most important transitions in the plant life cycle is from the vegetative to reproductive phase. During that transition, sucrose levels rise rapidly but transiently in the phloem and shoot apexes. For several species, the addition of exogenous sucrose promotes flowering, possibly by acting as a main signal. Although other sugars, including glucose, also appear to be involved in this transition, evidence for their roles in flowering is limited. In Arabidopsis thaliana, trehalose-6-phosphate serves as a signal to induce flowering. However, its roles in other plants have not been reported. Sucrose seems to function primarily in the leaf phloem to enhance the generation of florigens such as Flowering Locus T (FT) while trehalose-6-phosphate functions in the shoot apical meristem to promote the flowering signal pathway downstream of those florigens.