The transcription factor gene RDD4 contributes to the control of nutrient ion accumulation in rice.

In this study, we investigated the expression and functions of the transcription factor gene RDD4 (rice Dof daily fluctuations 4), which has sequence similarity to RDD1 that controls nutrient ion accumulation in rice. RDD4 protein was highly accumulated in leaf sheaths and localized to vascular bundles. RDD4-overexpressing plants (RDD4-OX) improved the accumulation of various nutrient ions, irrespective of nutrient concentration in a hydroponic solution. K+ and Cl- deficiencies induced the accumulation of other cations and anions, respectively. Interestingly, in RDD4-OX plants K+ and Cl- deficiencies increased PO4 3- and Mg2+ contents, respectively, despite opposite electric charges. Furthermore, PO4 3- deficiency induced NO3 - and Mg2+ accumulation in RDD4-OX plants. These data show that RDD4 is associated with the control of nutrient ion contents within plants. Also, photosynthetic CO2 assimilation in RDD4-OX plants was higher than in wild-type (WT) plants, although the sizes of shoots and panicles decreased in RDD4-OX plants. Subsequent microarray analysis indicated that OsFWL7, similar to maize CNR1 that negatively regulates plant size, showed the most significant difference in its expression levels between WT and RDD4-OX plants. Based on these results, it is hypothesized that a prominent increase in the OsFWL7 expression reduces plant size in RDD4-OX plants.

Arabidopsis ROOT PHOTOTROPISM2 Is a Light-Dependent Dynamic Modulator of Phototropin1.

The Arabidopsis (Arabidopsis thaliana) blue light photoreceptor phototropin1 (phot1) is a blue light-activated Ser/Thr protein kinase that mediates various light responses, including phototropism. The function of phot1 in hypocotyl phototropism is dependent on the light induction of ROOT PHOTOTROPISM2 (RPT2) proteins within a broad range of blue light intensities. It is not yet known however how RPT2 contributes to the photosensory adaptation of phot1 to high intensity blue light and the phototropic responses under bright light conditions. We show that RPT2 suppresses the activity of phot1 and demonstrate that RPT2 binds to the PHOT1 light, oxygen or voltage sensing1 (LOV1) domain that is required for its high photosensitivity. Our biochemical analyses revealed that RPT2 inhibits autophosphorylation of phot1, suggesting that it suppresses the photosensitivity and/or kinase activity of phot1 through the inhibition of LOV1 function. We found that RPT2 proteins are degraded via a ubiquitin-proteasome pathway when phot1 is inactive and are stabilized under blue light in a phot1-dependent manner. We propose that RPT2 is a molecular rheostat that maintains a moderate activation level of phot1 under any light intensity conditions.

Arabidopsis ROOT PHOTOTROPISM2 Contributes to the Adaptation to High-Intensity Light in Phototropic Responses.

Living organisms adapt to changing light environments via mechanisms that enhance photosensitivity under darkness and attenuate photosensitivity under bright light conditions. In hypocotyl phototropism, phototropin1 (phot1) blue light photoreceptors mediate both the pulse light-induced, first positive phototropism and the continuous light-induced, second positive phototropism, suggesting the existence of a mechanism that alters their photosensitivity. Here, we show that light induction of ROOT PHOTOTROPISM2 (RPT2) underlies photosensory adaptation in hypocotyl phototropism of Arabidopsis thaliana. rpt2 loss-of-function mutants exhibited increased photosensitivity to very low fluence blue light but were insensitive to low fluence blue light. Expression of RPT2 prior to phototropic stimulation in etiolated seedlings reduced photosensitivity during first positive phototropism and accelerated second positive phototropism. Our microscopy and biochemical analyses indicated that blue light irradiation causes dephosphorylation of NONPHOTOTROPIC HYPOCOTYL3 (NPH3) proteins and mediates their release from the plasma membrane. These phenomena correlate closely with the desensitization of phot1 signaling during the transition period from first positive phototropism to second positive phototropism. RPT2 modulated the phosphorylation of NPH3 and promoted reconstruction of the phot1-NPH3 complex on the plasma membrane. We conclude that photosensitivity is increased in the absence of RPT2 and that this results in the desensitization of phot1. Light-mediated induction of RPT2 then reduces the photosensitivity of phot1, which is required for second positive phototropism under bright light conditions.

OsNAC111, a blast disease-responsive transcription factor in rice, positively regulates the expression of defense-related genes.

Plants respond to pathogen attack by transcriptionally regulating defense-related genes via various types of transcription factors. We identified a transcription factor in rice, OsNAC111, belonging to the TERN subgroup of the NAC family that was transcriptionally upregulated after rice blast fungus (Magnaporthe oryzae) inoculation. OsNAC111 was localized in the nucleus of rice cells and had transcriptional activation activity in yeast and rice cells. Transgenic rice plants overexpressing OsNAC111 showed increased resistance to the rice blast fungus. In OsNAC111-overexpressing plants, the expression of several defense-related genes, including pathogenesis-related (PR) genes, was constitutively high compared with the control. These genes all showed blast disease-responsive expression in leaves. Among them, two chitinase genes and one ß-1,3-glucanase gene showed reduced expression in transgenic rice plants in which OsNAC111 function was suppressed by a chimeric repressor (OsNAC111-SRDX). OsNAC111 activated transcription from the promoters of the chitinase and ß-1,3-glucanase genes in rice cells. In addition, brown pigmentation at the infection sites, a defense response of rice cells to the blast fungus, was lowered in OsNAC111-SRDX plants at the early infection stage. These results indicate that OsNAC111 positively regulates the expression of a specific set of PR genes in the disease response and contributes to disease resistance.

Strigolactone and cytokinin act antagonistically in regulating rice mesocotyl elongation in darkness.

Strigolactones (SLs) are a group of phytohormones that control plant growth and development including shoot branching. Previous studies of the phenotypes of SL-related rice (Oryza sativa) dwarf (d) mutants demonstrated that SLs inhibit mesocotyl elongation by controlling cell division. Here, we found that the expression of cytokinin (CK)-responsive type-A RESPONSE REGULATOR (RR) genes was higher in d10-1 and d14-1 mutants than in the wild type. However, CK levels in mesocotyls of the d mutants were not very different from those in the wild type. On the other hand, application of a synthetic CK (kinetin) enhanced mesocotyl elongation in the d mutants and the wild type. d10-1 and d14-1 mesocotyls were more sensitive to CK than wild-type mesocotyls, suggesting that the up-regulation of the CK-responsive type-A RR genes and the higher elongation of mesocotyls in the d mutants are mainly due to the increased sensitivity of the d mutants to CK. Co-treatment with kinetin and a synthetic SL (GR24) confirmed the antagonistic functions of SL and CK on mesocotyl elongation. OsTCP5, which encodes a transcription factor belonging to the cell division-regulating TCP family, was also regulated by SL and CK and its expression was negatively correlated with mesocotyl length. These findings suggest that OsTCP5 contributes to the SL- and CK-controlled mesocotyl elongation in darkness.

Role of the phytochrome and cryptochrome signaling pathways in hypocotyl phototropism.

Unilateral blue-light irradiation activates phototropin (phot) photoreceptors, resulting in asymmetric distribution of the phytohormone auxin and induction of a phototropic response in higher plants. Other photoreceptors, including phytochrome (phy) and cryptochrome (cry), have been proposed as modulators of phototropic responses. We show here that either phy or cry is required for hypocotyl phototropism in Arabidopsis thaliana under high fluence rates of blue light, and that constitutive expression of ROOT PHOTOTROPISM 2 (RPT2) and treatment with the phytohormone gibberellin (GA) biosynthesis inhibitor paclobutrazol partially and independently complement the non-phototropic hypocotyl phenotype of the phyA cry1 cry2 mutant under high fluence rates of blue light. Our results indicate that induction of RPT2 and reduction in the GA are crucial for hypocotyl phototropic regulation by phy and cry. We also show that GA suppresses hypocotyl bending via destabilization of DELLA transcriptional regulators under darkness, but does not suppress the phototropic response in the presence of either phyA or cryptochromes, suggesting that these photoreceptors control not only the GA content but also the GA sensing and/or signaling that affects hypocotyl phototropism. The metabolic and signaling regulation of not only auxin but also GA by photoreceptors therefore appears to determine the hypocotyl growth pattern, including phototropic and gravitropic responses and inhibition of hypocotyl elongation, for adaptation to various light environments.

RPT2 is a signal transducer involved in phototropic response and stomatal opening by association with phototropin 1 in Arabidopsis thaliana.

Phototropin 1 (phot1) and phot2, which are blue light receptor kinases, function in blue light-induced hypocotyl phototropism, chloroplast relocation, and stomatal opening in Arabidopsis (Arabidopsis thaliana). Previous studies have shown that the proteins RPT2 (for ROOT PHOTOTROPISM2) and NPH3 (for NONPHOTOTROPIC HYPOCOTYL3) transduce signals downstream of phototropins to induce the phototropic response. However, the involvement of RPT2 and NPH3 in stomatal opening and in chloroplast relocation mediated by phot1 and phot2 was unknown. Genetic analysis of the rpt2 mutant and of a series of double mutants indicates that RPT2 is involved in the phot1-induced phototropic response and stomatal opening but not in chloroplast relocation or phot2-induced movements. Biochemical analyses indicate that RPT2 is purified in the crude microsomal fraction, as well as phot1 and NPH3, and that RPT2 makes a complex with phot1 in vivo. On the other hand, NPH3 is not necessary for stomatal opening or chloroplast relocation. Thus, these results suggest that phot1 and phot2 choose different signal transducers to induce three responses: phototropic response of hypocotyl, stomatal opening, and chloroplast relocation.

Isolation and expression analysis of petunia CURLY LEAF-like genes.

The Arabidopsis CURLY LEAF (CLF) gene is required to repress transcription of the class C gene AGAMOUS (AG) in whorls 1 and 2 of flowers and also in vegetative organs. CLF encodes a protein with homology to the product of the Drosophila Polycomb-group gene Enhancer of zeste [E(z)], which is involved in embyogenesis. In this study, we isolated three petunia CLF-like genes (PhCLF1, PhCLF2 and PhCLF3) based on the sequence homology between CLF and E(Z). Sequence analysis suggests that PhCLF1 and PhCLF2 are orthologs of CLF, whereas PhCLF3 is an ortholog of the Arabidopsis gene EZA1. We identified several conserved domains among products of PhCLF genes and related genes. PhCLF1 and PhCLF2 were expressed in all floral organs and leaves. The PhCLF1 transcripts were accumulated especially in corolla limbs, and contained several alternatively spliced RNA species. PhCLF1 and PhCLF2 do not appear to be the BLIND gene, which is required to repress transcription of the petunia class C gene, but their expression was affected by the homeotic conversion of organs in the blind flower. Our findings show that expression of PhCLF1 is regulated differently from that of PhCLF2, and suggest that the two petunia CLF orthologs function differently from each other.

Role of petunia pMADS3 in determination of floral organ and meristem identity, as revealed by its loss of function.

pMADS3, a petunia class C gene, is the candidate homologue of Arabidopsis AGAMOUS (AG), which is involved in the specification of stamens and carpels. We report the characterization of loss-of-function phenotype of pMADS3 that resulted from silencing of this gene. Silencing of pMADS3 resulted in homeotic conversion of stamens into petaloid structures, whereas the carpels were only weakly affected. Ectopic secondary inflorescences emerged from the interstamenal region in the third whorl, which is unique and has not been reported for any class C gene of other plant species. Third-order inflorescences emerged at corresponding positions in the third whorl of inner flowers of secondary inflorescences, indicating reiterative conversion of parts of the floral meristem into inflorescence meristem. On the basis of phenotypic analysis of the pMADS3-silenced plants, we propose that pMADS3 is involved in determination of floral organ and floral meristem identity in petunia. Two hybrid studies in yeast showed that PMADS3 protein interacted specifically with FBP2, a candidate homologue of Arabidopsis SEPALLATA3 (SEP3). The evidence presented here suggest that a complex involving PMADS3 and FBP2 is responsible for specification of organ identity in the third whorl.