Japonicus Roots Research Articles

Cross relationships of Cu, Fe, Zn, Mn, and Cd accumulations in common japonica and indica rice cultivars in Japan

Nineteen common rice cultivars in Japan (12 japonica and 7 indica) were grown on a basal media (basal) or that with either of excess but harmless amounts of Fe, Cu, Zn, or Cd (xsFe, xsCu, xsZn, and xsCd). Concentrations of these metals and Mn were then measured in shoots and roots to elucidate whether cross relationships of these metal accumulations differ in genotypes and/or tissues. Although each excess metal condition basically increased the administered metal both in shoots and roots irrespective of genotypes, the xsFe induced Fe-increase was not significant in japonica shoots. Also, xsFe increased Zn in indica shoots, and decreased Mn and Cu in indica roots and japonica shoots, respectively. Thus, xsFe induced all the observed alterations in metal accumulation, genotype- and/or tissue-specifically. On the other hand, inter-metal correlation coefficients could directly demonstrate effects of the excess metal conditions on individual sets of metal correlation. The xsFe induced Zn-increase and Cu-decrease more significantly affected the Zn/Mn correlation in indica shoots and the Cu/Fe correlation in japonica shoots, respectively. Interestingly, even though the effects of xsCd and xsZn were not obvious in the averaged metal concentration data, the correlation coefficients significantly indicated xsCd and xsZn induced changes in the Cu/Mn correlation in indica shoots and the Fe/Zn correlation in japonica roots, respectively. Furthermore, principal component analysis (PCA) could clearly visualize the difference in metal accumulation properties between shoots and roots and those between the japonica and indica genotypes. These results are the first demonstration of the genotype- and/or tissue-specific alterations in cross metal relationships, and serve as a basis for further analysis of the specificity in rice metal accumulations.

Atractylodes japonica root extract protects osteoblastic MC3T3‐E1 cells against hydrogen peroxide‐induced inhibition of osteoblastic differentiation

The rhizome of Atractylodes japonica Koidz. has been used traditionally for the treatment of water retention in the body and is known to have estrogen-like activity. In the present study, the protective effects of A. japonica root extract on the response of osteoblasts to oxidative stress were evaluated. Osteoblastic MC3T3-E1 cells were incubated with hydrogen peroxide and/or A. japonica root extract, and markers of osteoblast function and oxidative damage were examined. A. japonica root extract significantly (p < 0.05) increased cell survival, alkaline phosphatase activity, collagen and calcium deposition and decreased the production of receptor activator of nuclear factor-kB ligand (RANKL), protein carbonyl and malondialdehyde of osteoblastic MC3T3-E1 cells in the presence of hydrogen peroxide. These results demonstrate that A. japonica root can protect the cells from oxidative stress-induced toxicity and may have positive effects on skeletal structure.

Presymbiotic factors released by the arbuscular mycorrhizal fungus Gigaspora margarita induce starch accumulation in Lotus japonicus roots

* Nutrient exchange is the key symbiotic feature of arbuscular mycorrhiza (AM). As evidence is accumulating that plants sense presymbiotic factors from AM fungi and prepare for colonization, we investigated whether modifications in plant sugar metabolism might be part of the precolonization program. * Inoculation of Lotus japonicus roots in a double Millipore sandwich with the AM fungus Gigaspora margarita prevented contact between the symbionts but allowed exchange of signal molecules. Starch content was used as a marker for root carbohydrate status. * Mycorrhizal colonization of L. japonicus roots led to a decrease in starch concentration. In roots inoculated in the double sandwich, the polysaccharide accumulated after 1 wk and persisted for at least 4 wk. The response was absent in the castor myc(-) mutant, sym4-2, while transcript levels of both CASTOR and POLLUX were slightly enhanced in the wild-type L. japonicus roots, suggesting a requirement of the corresponding proteins for the starch-accumulation response. Exudates obtained from fungal spores germinated in the absence of the plant also induced starch accumulation in wild-type L. japonicus roots. * We conclude that factors released from germinating AM fungal spores induce changes in the root carbon status, possibly by enhancing sugar import, which leads to starch accumulation when colonization is prevented.

Multiple layers of incompatibility to the parasitic witchweed, Striga hermonthica

* Witchweeds (Striga spp.) are major agricultural pests that infest important crops in sub-Saharan Africa. Striga hermonthica parasitizes gramineous plants including sorghum, maize and rice, but not dicots. To understand host recognition mechanisms of S. hermonthica, we investigated its interaction with nonhost dicots including Arabidopsis, cowpea, Lotus japonicus and Phtheirospermum japonicum, a hemiparasite. * Striga hermonthica seeds were pretreated with strigol, a germination stimulant, and allowed to germinate next to a potential host root. We characterized the histological phenotype of the interactions. Moreover, we monitored the infection of a host rice and the nonhost P. japonicum by S. hermonthica using time-lapse photography. * All nonhost dicots tested did not support S. hermonthica shoot growth beyond the six leaf-pair stage; however, the arrest of parasite development occurred at different stages. Striga hermonthica haustoria were able to reach the steles of Arabidopsis and cowpea, while L. japonicus blocked S. hermonthica infection in the root cortex. Striga hermonthica often failed to penetrate P. japonicum roots. * Our analysis indicates that there are at least four types of incompatible interaction to S. hermonthica. Combinations of these different incompatibility mechanisms contribute to the total resistance to S. hermonthica.

A Mycorrhizal-Specific Ammonium Transporter from Lotus japonicus Acquires Nitrogen Released by Arbuscular Mycorrhizal Fungi

In mycorrhizal associations, the fungal partner assists its plant host by providing nitrogen (N) in addition to phosphate. Arbuscular mycorrhizal (AM) fungi have access to inorganic or organic forms of N and translocate them via arginine from the extra- to the intraradical mycelium, where the N is transferred to the plant without any carbon skeleton. However, the molecular form in which N is transferred, as well as the involved mechanisms, is still under debate. NH(4)(+) seems to be the preferential transferred molecule, but no plant ammonium transporter (AMT) has been identified so far. Here, we offer evidence of a plant AMT that is involved in N uptake during mycorrhiza symbiosis. The gene LjAMT2;2, which has been shown to be the highest up-regulated gene in a transcriptomic analysis of Lotus japonicus roots upon colonization with Gigaspora margarita, has been characterized as a high-affinity AMT belonging to the AMT2 subfamily. It is exclusively expressed in the mycorrhizal roots, but not in the nodules, and transcripts have preferentially been located in the arbusculated cells. Yeast (Saccharomyces cerevisiae) mutant complementation has confirmed its functionality and revealed its dependency on acidic pH. The transport experiments using Xenopus laevis oocytes indicated that, unlike other plant AMTs, LjAMT2;2 transports NH(3) instead of NH(4)(+). Our results suggest that the transporter binds charged ammonium in the apoplastic interfacial compartment and releases the uncharged NH(3) into the plant cytoplasm. The implications of such a finding are discussed in the context of AM functioning and plant phosphorus uptake.

Rearrangement of Actin Cytoskeleton Mediates Invasion ofLotus japonicusRoots byMesorhizobium loti

Infection thread-dependent invasion of legume roots by rhizobia leads to internalization of bacteria into the plant cells, which is one of the salient features of root nodule symbiosis. We found that two genes, Nap1 (for Nck-associated protein 1) and Pir1 (for 121F-specific p53 inducible RNA), involved in actin rearrangements were essential for infection thread formation and colonization of Lotus japonicus roots by its natural microsymbiont, Mesorhizobium loti. nap1 and pir1 mutants developed an excess of uncolonized nodule primordia, indicating that these two genes were not essential for the initiation of nodule organogenesis per se. However, both the formation and subsequent progression of infection threads into the root cortex were significantly impaired in these mutants. We demonstrate that these infection defects were due to disturbed actin cytoskeleton organization. Short root hairs of the mutants had mostly transverse or web-like actin filaments, while bundles of actin filaments in wild-type root hairs were predominantly longitudinal. Corroborating these observations, temporal and spatial differences in actin filament organization between wild-type and mutant root hairs were also observed after Nod factor treatment, while calcium influx and spiking appeared unperturbed. Together with various effects on plant growth and seed formation, the nap1 and pir1 alleles also conferred a characteristic distorted trichome phenotype, suggesting a more general role for Nap1 and Pir1 in processes establishing cell polarity or polar growth in L. japonicus.

オウレン抽出物とその主成分ベルベリンがヒト皮下由来脂肪細胞に及ぼす分化誘導抑制作用および脂肪分解促進作用

オウレン抽出物とその主成分ベルベリンがヒト皮下由来脂肪細胞に及ぼす分化誘導抑制作用および脂肪分解促進作用

Reactions of Lotus japonicus ecotypes and mutants to root parasitic plants

Witchweeds (Striga spp.) and broomrapes (Orobanche spp.) are obligate root parasitic plants on economically important field and horticultural crops. The parasites' seeds are induced to germinate by root-derived chemical signals. The radicular end is transformed into a haustorium which attaches, penetrates the host root and establishes connection with the vascular system of the host. Reactions of Lotus japonicus, a model legume for functional genomics, were studied for furthering the understanding of host-parasite interactions. Lotus japonicus was compatible with Orobanche aegyptiaca, but not with Orobanche minor, Striga hermonthica and Striga gesnerioides. Orobanche minor successfully penetrated Lotus japonicus roots, but failed to establish connections with the vascular system. Haustoria in Striga hermonthica attached to the roots, but penetration and subsequent growth of the endophyte in the cortex were restricted. Striga gesnerioides did not parasitize Lotus japonicus. Among seven mutants of Lotus japonicus (castor-5, har1-5, alb1-1, ccamk-3, nup85-3, nfr1-3 and nsp2-1) with altered characteristics in relation to rhizobial nodulation and mycorrhizal colonization, castor-5 and har1-5 were parasitized by Orobanche aegyptiaca with higher frequency than the wild type. In contrast, Orobanche aegyptiaca tubercle development was delayed on the mutants nup85-3, nfr1-3 and nsp2-1. These results suggest that nodulation, mycorrhizal colonization and infection by root parasitic plants in Lotus japonicus may be modulated by similar mechanisms and that Lotus japonicus is a potential model legume for studying plant-plant parasitism.

Anti-tumor actions of major component 3′- O-acetylhamaudol of Angelica japonica roots through dual actions, anti-angiogenesis and intestinal intraepithelial lymphocyte activation

We recently demonstrated that two chalcones isolated from Angelica keiskei roots have anti-tumor and anti-metastatic activities through the inhibition of tumor-induced angiogenesis, but the anti-tumor substances of Angelica japonica roots are unknown. We attempted to clarify the anti-tumor action and its mechanisms of a major component 3′- O-acetylhamaudol isolated from A. japonica roots. We first examined the effects of 3′- O-acetylhamaudol on tumor growth in colon 26-bearing mice. Furthermore, we examined the effects of 3′- O-acetylhamaudol on angiogenic factors (vascular endothelial growth factor receptor-2 (VEGFR-2) phosphorylation in human umbilical vein endothelial cells (HUVECs), and vascular endothelial growth factor (VEGF) production and hypoxia-inducible factor (HIF)-1α expression in tumors). 3′- O-Acetylhamaudol (25 and 50 mg/kg, twice daily) inhibited the tumor growth in colon 26-bearing mice. Although 3′- O-acetylhamudol had no effect on the VEGF production and HIF-1α in colon 26 cells, it (10 μM) inhibited the VEGF-induced angiogenesis and VEGF-induced VEGFR-2 phosphorylation in HUVECs. 3′- O-Acetylhamaudol has anti-tumor effects mediated through dual mechanisms, i.e., anti-angiogenic actions and the modulation of the immune system in the spleen and small intestine in tumor-bearing mice.

13. Induction of phytoalexin biosynthesis in Lotus japonicus roots in response to Striga hermonthica attachment

13. Induction of phytoalexin biosynthesis in Lotus japonicus roots in response to Striga hermonthica attachment

Defects in Rhizobial Cyclic Glucan and Lipopolysaccharide Synthesis Alter Legume Gene Expression During Nodule Development

cDNA array technology was used to compare transcriptome profiles of Lotus japonicus roots inoculated with a Mesorhizobium loti wild-type and two mutant strains affected in cyclic beta(1-2) glucan synthesis (cgs) and in lipopolysaccharide synthesis (lpsbeta2). Expression of genes associated with the development of a fully functional nodule was significantly affected in plants inoculated with the cgs mutant. Array results also revealed that induction of marker genes for nodule development was delayed when plants were inoculated with the lpsbeta2 mutant. Quantitative real-time reverse-transcriptase polymerase chain reaction was used to quantify gene expression of a subset of genes involved in plant defense response, redox metabolism, or genes that encode for nodulins. The majority of the genes analyzed in this study were more highly expressed in roots inoculated with the wild type compared with those inoculated with the cgs mutant strain. Some of the genes exhibited a transient increase in transcript levels during intermediate steps of normal nodule development while others displayed induced expression during the final steps of nodule development. Ineffective nodules induced by the glucan mutant showed higher expression of phenylalanine ammonia lyase than wild-type nodules. Differences in expression pattern of genes involved in early recognition and signaling were observed in plants inoculated with the M. loti mutant strain affected in the synthesis of cyclic glucan.

Antioxidant and antidiabetic activities of extracts from Cirsium japonicum roots

This study investigated the antioxidant activity of methanol (MeOH) and water extracts from roots of Cirsium japonicum in vitro. MeOH extract showed a stronger free radical scavenging activity than water extract. However, both of extracts showed a concentration dependent hydroxyl radical scavenging activity, reducing power and metal chelating ability. MeOH extract had greater phenolic and flavonoid contents than water extract. The antidiabetic activity of these two extracts was evaluated by the α-glucosidase inhibition assay. The water extract showed a considerable α-glucosidase inhibitory activity. To our knowledge, this may be the first time to report the antioxidant and antidiabetic activities in Cirsium japonicum roots.

A Mesorhizobium loti mutant with reduced glucan content shows defective invasion of its host plant Lotus japonicus

Random transposon mutagenesis led to the isolation of a novel Mesorhizobium loti mutant that is defective in nitrogen fixation during symbiosis with Lotus japonicus. The mutated locus, designated cep, encodes a putative cell-envelope protein displaying no significant sequence similarity to proteins with known functions. This mutant elicits the formation of nodule-like bumps and root-hair curling, but not the elongation of infection threads, on L. japonicus roots. This is reminiscent of the phenotypes of rhizobial mutants impaired in cyclic beta-glucan biosynthesis. The cep mutant exhibits partially reduced content of cell-associated glucans and intermediate deficiency of motility under hypo-osmotic conditions as compared to a glucan-deficient mutant. Second-site pseudorevertants of the cep mutant were isolated by selecting for restoration of symbiotic nitrogen fixation. A subset of pseudorevertants restored both symbiotic capability and glucan content to levels comparable to that of the wild-type. These results suggest that the Cep product acts on a successful symbiosis by affecting cell-associated glucan content.

Production of (+)-5-deoxystrigol by Lotus japonicus root culture

Lotus japonicus roots, cultured in a modified B5 medium, produced and secreted germination stimulants that induced Striga hermonthica seed germination. The germination-inducing activity was detected both in the roots and the culture filtrate. Following bioassay-guided purification procedures, an active compound was isolated from hexane extracts of the roots and the culture filtrate. Based on chromatographic behaviour on HPLC, and 1H NMR, UV, MS and CD spectroscopic analyses, the germination stimulant was identified as (+)-5-deoxystrigol.

Transcriptome Profiling of Lotus japonicus Roots During Arbuscular Mycorrhiza Development and Comparison with that of Nodulation

To better understand the molecular responses of plants to arbuscular mycorrhizal (AM) fungi, we analyzed the differential gene expression patterns of Lotus japonicus, a model legume, with the aid of a large-scale cDNA macroarray. Experiments were carried out considering the effects of contaminating microorganisms in the soil inoculants. When the colonization by AM fungi, i.e. Glomus mosseae and Gigaspora margarita, was well established, four cysteine protease genes were induced. In situ hybridization revealed that these cysteine protease genes were specifically expressed in arbuscule-containing inner cortical cells of AM roots. On the other hand, phenylpropanoid biosynthesis-related genes for phenylalanine ammonia-lyase (PAL), chalcone synthase, etc. were repressed in the later stage, although they were moderately up-regulated on the initial association with the AM fungus. Real-time RT–PCR experiments supported the array experiments. To further confirm the characteristic expression, a PAL promoter was fused with a reporter gene and introduced into L. japonicus, and then the transformants were grown with a commercial inoculum of G. mosseae. The reporter activity was augmented throughout the roots due to the presence of contaminating microorganisms in the inoculum. Interestingly, G. mosseae only colonized where the reporter activity was low. Comparison of the transcriptome profiles of AM roots and nitrogen-fixing root nodules formed with Mesorhizobium loti indicated that the PAL genes and other phenylpropanoid biosynthesis-related genes were similarly repressed in the two organs.

Pore space organization and plant response in peat substrates: I. Prunus x cistena and Spiraea japonica

Aeration properties are key characteristics of growing media. On top of air filled porosity, pore tortuosity (t) and gas diffusivity (Ds/Do) have been suggested to better characterize the aeration process. A two-year nursery study was conducted to investigate how Ds/Do of growing mediawere affected by particle size and how plant growth responded to substrates with the same air storage, but different gas exchange properties. Four substrates were made of the same proportion (on a volume basis) of sphagnum peat (50%), composted coniferous wood bark (40%) and gravel (10%). Different particle sizes of coniferous bark (1 - 2, 2 - 4, 4 - 8 and 8 - 16 mm) were used. However, air-filled porosity (¦a) remained unaffected by this change in bark particle size. The factor t increased linearly with increasing bark particle size while Ds/Dodecreased. Such substrates were used to grow Prunus cystena sp and Spirae japonica for two years in 5 L pot, at three different water potentials (-0.8, -1.2, -1.6 kPa) Increasing coniferous wood bark particle size decreased plant growth significantly for nearly all parameters studied. Prunus x cistena sp and Spiraea japonica shoot and root dry weights were very significantly correlated with both t and Ds/Do with determination coefficients ranging from 0.43 to 0.71. This study suggests a causal-effect relationship between plant growth and substrate gas exchange characteristics. Key words: Gas diffusivity, pore tortuosity, air-filled porosity, peat substrates, peat-lite mixes,Prunus x cistena, Spiraea japonica.

Grafting between model legumes demonstrates roles for roots and shoots in determining nodule type and host/rhizobia specificity

Previous grafting experiments have demonstrated that legume shoots play a critical role in symbiotic development of nitrogen-fixing root nodules by regulating nodule number. Here, reciprocal grafting experiments between the model legumes Lotus japonicus and Medicago truncatula were carried out to investigate the role of the shoot in the host-specificity of legume-rhizobia symbiosis and nodule type. Lotus japonicus is nodulated by Mesorhizobium loti and makes determinate nodules, whereas M. truncatula is nodulated by Sinorhizobium meliloti and makes indeterminate nodules. When inoculated with M. loti, L. japonicus roots grafted on M. truncatula shoots produced determinate nodules identical in appearance to those produced on L. japonicus self-grafted roots. Moreover, the hypernodulation phenotype of L. japonicus har1-1 roots grafted on wild-type M. truncatula shoots was restored to wild type when nodulated with M. loti. Thus, L. japonicus shoots appeared to be interchangeable with M. truncatula shoots in the L. japonicus root/M. loti symbiosis. However, M. truncatula roots grafted on L. japonicus shoots failed to induce nodules after inoculation with S. meliloti or a mixture of S. meliloti and M. loti. Instead, only early responses to S. meliloti such as root hair tip swelling and deformation, plus induction of the early nodulation reporter gene MtENOD11:GUS were observed. The results indicate that the L. japonicus shoot does not support normal symbiosis between the M. truncatula root and its microsymbiont S. meliloti, suggesting that an unidentified shoot-derived factor may be required for symbiotic progression in indeterminate nodules.

Invasion of Lotus japonicus root hairless 1 by Mesorhizobium loti Involves the Nodulation Factor-Dependent Induction of Root Hairs

In many legumes, including Lotus japonicus and Medicago truncatula, susceptible root hairs are the primary sites for the initial signal perception and physical contact between the host plant and the compatible nitrogen-fixing bacteria that leads to the initiation of root invasion and nodule organogenesis. However, diverse mechanisms of nodulation have been described in a variety of legume species that do not rely on root hairs. To clarify the significance of root hairs during the L. japonicus-Mesorhizobium loti symbiosis, we have isolated and performed a detailed analysis of four independent L. japonicus root hair developmental mutants. We show that although important for the efficient colonization of roots, the presence of wild-type root hairs is not required for the initiation of nodule primordia (NP) organogenesis and the colonization of the nodule structures. In the genetic background of the L. japonicus root hairless 1 mutant, the nodulation factor-dependent formation of NP provides the structural basis for alternative modes of invasion by M. loti. Surprisingly, one mode of root colonization involves nodulation factor-dependent induction of NP-associated cortical root hairs and epidermal root hairs, which, in turn, support bacterial invasion. In addition, entry of M. loti through cracks at the cortical surface of the NP is described. These novel mechanisms of nodule colonization by M. loti explain the fully functional, albeit significantly delayed, nodulation phenotype of the L. japonicus ROOT HAIRLESS mutant.

Coptis japonica root extract induces apoptosis through caspase3 activation in SNU-668 human gastric cancer cells

Apoptosis-modulating approaches offer an attractive opportunity for therapeutic use for many tumors. We investigated the effects of the roots of Coptis japonica var. dissecta (Ranunculaceae) on human gastric cancer cells, SNU-668. The cytotoxicity of Coptis japonica at 100 microg/ml (methanol extract) by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay was 13.89 +/- 1.91% of control value. Considering the features by 4,6-diamidino-2-phenylindole (DAPI) staining and terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay, it was confirmed that the death of SNU-668 cells was due to apoptosis. In the apoptosis-regulating genes, BCL2 expression was diminished out, whereas BAX and CASP3 expressions were increased, compared with control. Furthermore, the activity of caspase3 was significantly increased by Coptis japonica treatment. These results suggest that Coptis japonica could induce apoptotic anticancer effect through caspase3 activation on SNU-668 human gastric cancer cells.

Plant growth inhibitory activity of Ophiopogon japonicus Ker-Gawler and role of phenolic acids and their analogues: a comparative study

Allelopathic potential of Ophiopogon japonicus was investigated. The methanolic extract of O. japonicus roots strongly inhibited root and hypocotyls growth of lettuce. Sequential partitioning of the methanol extract with organic solvents showed that the diethyl ether and n-butanol extract possess strong plant growth inhibitory activities. The allelopathic constituents of the diethyl ether extract were isolated and identified as salicylic acid and p-hydroxybenzoic acid by NMR spectroscopy. Both of these phenolic acids were found in the aqueous extracts of leaves as well. The concentration of salicylic acid in roots and leaves were estimated as 0.011 and 0.02%, respectively, and it inhibited the root and shoot of tested plants by 50% even at less than 3 ppm. The p-hydroxybenzoic acid on the other hand was in less abundance (0.005%) and inhibited the plant growth to a lesser extent. The biological activity of commercially available O-methyl derivatives of these phenolic acids was also determined to establish structure–activity relationship. Among these, salicylic acid was found to be the most active one. These results suggest that Ophiopogon japonicus produces plant growth inhibitors, which are responsible for its potential allelopathic activity.