Expression and functions of myo-inositol monophosphatase family genes in seed development of Arabidopsis.

Myo-inositol monophosphatase (IMP) catalyzes the dephosphorylation of myo-inositol 3-phosphate in the last step of myo-inositol biosynthesis. IMP is also important in phosphate metabolism and is required for the biosynthesis of cell wall polysaccharides, phytic acid, and phosphatidylinositol. In Arabidopsis, IMP is encoded by VTC4. There are, however, two additional IMP candidate genes, IMPL1 and IMPL2, which have not yet been elucidated. In our genetic studies of Arabidopsis IMP genes, only the loss-of-function mutant impl2 showed embryonic lethality at the globular stage. All IMP genes were expressed in a similar manner both in the vegetative and reproductive organs. In developing seeds, expression of IMP genes was not coupled with the expression of the genes encoding myo-inositol phosphate synthases, which supply the substrate for IMPs in the de novo synthesis pathway. Instead, expression of IMP genes was correlated with expression of the gene for myo-inositol polyphosphate 1-phosphatase (SAL1), which is involved in the myo-inositol salvage pathway, suggesting a possible salvage pathway role in seed development. Moreover, the partial rescue of the impl2 phenotype by histidine application implies that IMPL2 is also involved in histidine biosynthesis during embryo development.

Role of OsNPR1 in rice defense program as revealed by genome-wide expression analysis.

NPR1 is a central regulator of salicylic-acid (SA)-mediated defense signaling in Arabidopsis. Here, we report the characterization of OsNPR1, an Oryzae sativa (rice) ortholog of NPR1, focusing on its role in blast disease resistance and identification of OsNPR1-regulated genes. Blast resistance tests using OsNPR1 knockdown and overexpressing rice lines demonstrated the essential role of OsNPR1 in benzothiadiazole (BTH)-induced blast resistance. Genome-wide transcript profiling using OsNPR1-knockdown lines revealed that 358 genes out of 1,228 BTH-upregulated genes and 724 genes out of 1,069 BTH-downregulated genes were OsNPR1-dependent with respect to BTH responsiveness, thereby indicating that OsNPR1 plays a more vital role in gene downregulation. The OsNPR1-dependently downregulated genes included many of those involved in photosynthesis and in chloroplast translation and transcription. Reduction of photosynthetic activity after BTH treatment and its negation by OsNPR1 knockdown were indeed reflected in the changes in Fv/Fm values in leaves. These results imply the role of OsNPR1 in the reallocation of energy and resources during defense responses. We also examined the OsNPR1-dependence of SA-mediated suppression of ABA-induced genes.

Abscisic acid interacts antagonistically with salicylic acid signaling pathway in rice-Magnaporthe grisea interaction.

Plant hormones play pivotal signaling roles in plant-pathogen interactions. Here, we report characterization of an antagonistic interaction of abscisic acid (ABA) with salicylic acid (SA) signaling pathways in the rice-Magnaporthe grisea interaction. Exogenous application of ABA drastically compromised the rice resistance to both compatible and incompatible M. grisea strains, indicating that ABA negatively regulates both basal and resistance gene-mediated blast resistance. ABA markedly suppressed the transcriptional upregulation of WRKY45 and OsNPR1, the two key components of the SA signaling pathway in rice, induced by SA or benzothiadiazole or by blast infection. Overexpression of OsNPR1 or WRKY45 largely negated the enhancement of blast susceptibility by ABA, suggesting that ABA acts upstream of WRKY45 and OsNPR1 in the rice SA pathway. ABA-responsive genes were induced during blast infection in a pattern reciprocal to those of WRKY45 and OsPR1b in the compatible rice-blast interaction but only marginally in the incompatible one. These results suggest that the balance of SA and ABA signaling is an important determinant for the outcome of the rice-M. grisea interaction. ABA was detected in hyphae and conidia of M. grisea as well as in culture media, implying that blast-fungus-derived ABA could play a role in triggering ABA signaling at host infection sites.

Large-scale analysis of full-length cDNAs from the tomato (Solanum lycopersicum) cultivar Micro-Tom, a reference system for the Solanaceae genomics.

BACKGROUND: The Solanaceae family includes several economically important vegetable crops. The tomato (Solanum lycopersicum) is regarded as a model plant of the Solanaceae family. Recently, a number of tomato resources have been developed in parallel with the ongoing tomato genome sequencing project. In particular, a miniature cultivar, Micro-Tom, is regarded as a model system in tomato genomics, and a number of genomics resources in the Micro-Tom-background, such as ESTs and mutagenized lines, have been established by an international alliance. RESULTS: To accelerate the progress in tomato genomics, we developed a collection of fully-sequenced 13,227 Micro-Tom full-length cDNAs. By checking redundant sequences, coding sequences, and chimeric sequences, a set of 11,502 non-redundant full-length cDNAs (nrFLcDNAs) was generated. Analysis of untranslated regions demonstrated that tomato has longer 5'- and 3'-untranslated regions than most other plants but rice. Classification of functions of proteins predicted from the coding sequences demonstrated that nrFLcDNAs covered a broad range of functions. A comparison of nrFLcDNAs with genes of sixteen plants facilitated the identification of tomato genes that are not found in other plants, most of which did not have known protein domains. Mapping of the nrFLcDNAs onto currently available tomato genome sequences facilitated prediction of exon-intron structure. Introns of tomato genes were longer than those of Arabidopsis and rice. According to a comparison of exon sequences between the nrFLcDNAs and the tomato genome sequences, the frequency of nucleotide mismatch in exons between Micro-Tom and the genome-sequencing cultivar (Heinz 1706) was estimated to be 0.061%. CONCLUSION: The collection of Micro-Tom nrFLcDNAs generated in this study will serve as a valuable genomic tool for plant biologists to bridge the gap between basic and applied studies. The nrFLcDNA sequences will help annotation of the tomato whole-genome sequence and aid in tomato functional genomics and molecular breeding. Full-length cDNA sequences and their annotations are provided in the database KaFTom http://www.pgb.kazusa.or.jp/kaftom/ via the website of the National Bioresource Project Tomato http://tomato.nbrp.jp.

Possible involvement of DNA methylation on expression regulation of carrot LEC1 gene in its 5'-upstream region.

DNA methylation plays important roles in various developmental processes in many organisms. In carrots, the treatment of embryogenic cells (ECs) with DNA methylation inhibitors induces hypomethylation and blocks somatic embryogenesis. CARROT-LEAFY COTYLEDON 1 (C-LEC1) is an important transcription factor for embryo development that shows embryo-specific expression in ECs and somatic and zygotic embryos. However, the regulation of embryo-specific transcription factor genes such as C-LEC1 in plants is not well understood. In this study, we used embryogenic carrot cells (Daucus carota L. cv. US-Harumakigosun) to investigate the DNA methylation status of the embryogenesis-related genes C-LEC1, Carrot ABA INSENSITIVE 3 (C-ABI3), and Daucus carota Embryogenic cell protein 31 (DcECP 31) during the transition from embryogenesis to vegetative growth. The C-LEC1 promoter region showed a reduced level of DNA methylation during somatic embryogenesis followed by an increase during the transition from embryonic to vegetative growth. To test whether the increased level of DNA methylation down-regulates C-LEC1 expression, RNA-directed DNA methylation (RdDM) was used to induce the hypermethylation of two segments of the C-LEC1 5'-upstream region: Regions 1 and 2, corresponding to nucleotides -1,904 to -1,272 and -896 to -251, respectively. When the hypermethylation of Region 1 was induced by RdDM, C-LEC1 expression was reduced in the transgenic ECs, indicating a negative correlation between DNA methylation and C-LEC1 expression. In contrast, the hypermethylation of Region 2 did not greatly affect C-LEC1 expression. Based on these results, we hypothesize that DNA methylation may be involved in the control of C-LEC1 expression during carrot embryogenesis.

Identification and characterization of carrot HAP factors that form a complex with the embryo-specific transcription factor C-LEC1.

C-LEC1, an orthologue of Arabidopsis LEC1, is thought to be an essential transcriptional activator required for normal development during the early and late phases of embryogenesis. C-LEC1 is similar in sequence to the HAP3 subunits of other organisms. To understand C-LEC1 function better, a cDNA library of carrot somatic embryos was screened for factors that form complexes with C-LEC1. Two carrot HAP5 homologues and two carrot HAP2 homologues were identified; these factors have significant sequence similarity to the conserved regions of HAP5 and HAP2, respectively. Some of these proteins form heterotrimeric complexes that bind specifically to DNA fragments containing a CCAAT sequence in vitro. The results suggest that C-LEC1 is a component of the CCAAT-box-binding factor and forms a complex with C-HAP2B and C-HAP5A or C-HAP5B that regulates gene expression during carrot embryo development.

Isolation of the gene encoding Carrot leafy cotyledon1 and expression analysis during somatic and zygotic embryogenesis.

The Arabidopsis thaliana LEC1 gene regulates embryo morphology and seed maturation. For a better understanding of its function, we isolated a carrot (Daucus carota L. cv. US-Harumakigosun) counterpart of this gene, C-LEC1, from a cDNA library of carrot somatic embryos, since carrot is a better model plant for preparing large quantities of somatic embryos at the same developmental stage. The predicted amino acid sequence of C-LEC1 is similar to that of LEC1 and contains regions that are conserved in the heme-activated protein 3 (HAP3) subunit of plants, animals and microorganisms. C-LEC1 expression was detected in embryogenic cells, somatic embryos, and developing seeds. In situ hybridization analysis revealed C-LEC1 expression in the peripheral region of the embryos but not in the endosperm. Expression of C-LEC1 driven by Arabidopsis LEC1 promoter was able to complement the defects of the Arabidopsis lec1-1 mutant. These results suggest that C-LEC1 is a functional homolog of Arabidopsis LEC1, an important regulator of zygotic and somatic embryo development.