Statistical interferometric technique (SIT) is a highly sensitive, high speed non-contact, and non-destructive optical technique developed by our group capable of measuring instantaeoues sub-nanometer displacements. SIT applied to plant leaf elongation revealed nanometric intrinsic fluctuaitons (NIF) that are robust and sensitive to variations in the environment making NIF as a measure of healthiness of the plants. In this study, exogenous plant hormones, auxin (2,4-dichlorophenoxyacetic acid-2,4-D), and gibberellic acid (GA3), along with an auxin transport inhibitor 2,3,5-triiodobenzoic acid-TIBA, that affect plant growth were used to investigate their effects on NIF. Rice (Oriza sativa) seedlings were used, and their roots were exposed to 1, 2, and 4 µM 2,4-D, and the auxin transport inhibitor, TIBA, of 10, and 20 µM for 22 h and GA3 solution of different concentrations of 10, 40, and 100 µM for 5 h. Results showed significant increment in NIF for 1 µM and reduction for 4 µM 2,4-D while applicaiton of both 10, and 20 µM TIBA led to reduction in NIF. On the other hand, significant increment in NIF for 40 µM, and a significant reduction at a higher concentration of 100 µM for 5 hours of GA3 were also observed in comparison to those of control. Our results indicate that NIF as revealed by SIT could show both the positive and negative effects depending on the concentration of exogenous hormones, and transport inhibitors. Results suggest that SIT could be a valuable tool being sensitive enough to speedily assess the effects of plant growth hormones.
The root-knot nematode (RKN) Meloidogyne incognita is one of the most economically damaging plant-parasitic nematodes. Molecular studies of the plant–RKN interaction have been vigorously carried out in dicotyledonous model plants, while the host range of M. incognita is wide including monocotyledonous plants. As M. incognita causes quality and yield losses in rice (Oryza sativa L.) cultivated in both upland and irrigated systems, we developed a method to examine the plant–RKN interaction in this model monocotyledonous crop plant. Here, we show that a transparent paper pouch could be used to evaluate nematode infection rates in rice with similar results to that of the traditional soil method. The system using a transparent paper pouch can be used to observe the spatial and temporal distribution of developing galls and can save the space of growth chamber.
Phosphate (Pi) starvation affects root hair formation to increase the absorptive surface area of the roots. CAPRICE (CPC) and its homolog genes, including TRIPTYCHON (TRY), ENHANCER OF TRY AND CPC1 (ETC1), ETC2, and ETC3, positively regulate root hair formation in a partially redundant manner. In particular, ETC1 responds to Pi deficiency. To clarify role sharing among the CPC homolog genes under Pi-deficient condition, we analyzed the expression of five CPC homolog genes under Pi-deficient condition, using the real-time polymerase chain reaction analysis. Pi starvation enhanced the expression of not only ETC1, but also ETC3. Furthermore, ETC3, which is rarely expressed in the roots, was induced by Pi deficiency. The expression levels of CPC, TRY, and ETC2 in response to Pi deficiency were not significantly different from those under the control conditions. These results suggest that CPC homologs can be divided into two groups, genes that respond to Pi deficiency (ETC1 and ETC3) and those that do not (CPC, TRY, and ETC2).
Anthraquinones are widely distributed in various organisms and known as bioactive ingredients. Some of the anthraquinones accumulate as glycosides in higher plants. Plant secondary product glycosyltransferases (PSPGs) are the well-characterized enzymes producing plant secondary metabolite glycosides. However, PSPGs involved in the formation of anthraquinone glycosides remains unclear. The rhizome of Rheum palmatum contains anthraquinones as laxative agents, some of which are accumulated as glucosides. We isolated a glucosyltransferase, R. palmatum UDP-glycosyltransferase (RpUGT) 1 from the rhizome of R. palmatum, and characterized functionally. RpUGT1 glucosylated emodin yielding emodin-6-O-glucoside, and it also glucosylated rhapontigenin, a compound belonging to stilbenes, yielding rhaponticin. The expression patterns of RpUGT1 and the accumulation of the metabolites revealed that RpUGT1 contributes to the production of these glucosides in R. palmatum. These results may provide important information for the substrate recognition of the PSPGs for anthraquinones and stilbenes.
Most leguminous plants produce (−)-type enantiomers of pterocarpans as the phytoalexin, but pea (Pisum sativum L.) produces the opposite stereoisomer of pterocarpan, (+)-pisatin. Biosynthesis of (−)-pterocarpan skeleton is completely characterized at the molecular level, and pterocarpan synthase (PTS), a dirigent (DIR) domain-containing protein, participates in the last dehydration reaction. Similarly, isoflav-3-ene, a precursor of (+)-pisatin, is likely to be biosynthesized by the DIR-mediated dehydration reaction; however the biosynthesis is still unknown. In the present study, we screened PTS homologs based on RNA-sequence data from (+)-pisatin-producing pea seedlings and demonstrated that one of the candidates encodes isoflav-3-ene synthase (I3S). Real-time PCR analysis revealed that transcripts of I3S, in addition to other genes involved in the (+)-pisatin pathway, transiently accumulated in pea upon elicitation prior to the maximum accumulation of (+)-pisatin. I3S orthologs were also found in soybean and Lotus japonicus that are not known to accumulate (+)-pterocarpan, and the catalytic function of gene products was verified to be I3S by the in vitro enzyme assay. Incubation of the crude extract of elicited soybean cells with isoflav-3-ene yielded coumestrol, suggesting that isoflav-3-ene is a precursor of coumestrol biosynthesis in soybean.
Lipocalins are very important proteins for stress resistance in plants. To better understand the function of tomato lipocalins, we observed responses to oxidative stress using over-expressed SlTIL1, SlTIL2, SlCHL, and silenced-plants. Significant differences in reactive oxygen species accumulation (oxidative damage) were observed in all tested plants under heat stress. Plants with over-expressed SlTIL1, SlTIL2, and SlCHL showed less oxidative damage compared with wild-type plants under heat stress. The expression of SlSODs was induced in over-expressed SlTIL1, SlTIL2, and SlCHL plants under normal and heat stress conditions. Furthermore, silenced PDS, SlTILs, and SlCHL plants showed slightly increasing oxidative damage under heat stress alongside with lower SlSODs under normal and stress conditions. These results suggest that SlTIL1, SlTIL2, and SlCHL were involved in antioxidant defense by eliminating ROS in tomato plants.
Receptor complex formation at the cell surface is a key step to initiate downstream signaling but the contribution of this process for the regulation of the direction of downstream responses is not well understood. In the plant-microbe interactions, while CERK1, an Arabidopsis LysM-RLK, mediates chitin-induced immune responses, NFR1, a Lotus homolog of CERK1, regulates the symbiotic process with rhizobial bacteria through the recognition of Nod factors. Concerning the mechanistic insight of the regulation of such apparently opposite biological responses by the structurally related RLKs, Nakagawa et al. previously showed that the addition of YAQ sequence, conserved in NFR1 and other symbiotic LysM-RLKs, to the kinase domain of CERK1 switched downstream responses from defense to symbiosis using a set of chimeric receptors, NFR1-CERK1s. These results indicated that such a subtle difference in the cytoplasmic domain of LysM-RLKs could determine the direction of host responses from defense to symbiosis. On the other hand, it is still not understood how such structural differences in the cytoplasmic domains determine the direction of host responses. We here analyzed the interaction between chimeric NFR1s and NFR5, a partner receptor of NFR1, by co-immunoprecipitation (Co-IP) of these proteins transiently expressed in Nicotiana benthamiana. These results indicated that the cytoplasmic interaction between the LysM-RLKs is important for the symbiotic receptor complex formation and the YAQ containing region of NFR1 contributes to trigger symbiotic signaling through the successful formation of NFR1/NFR5 complex.
Phosphatidic acid plays an important role in plant immune responses against phytopathogenic bacteria in Nicotiana benthamiana. Here we focused on phosphoinositide dependent protein kinases (PDKs) as a candidate required for phosphatidic acid signaling. Based on Arabidopsis PDK sequences, we identified four putative PDK orthologs in N. benthamiana genome. To address the role of PDKs in plant defense responses, we created all four NbPDKs-silenced plants by virus-induced gene silencing. the NbPDKs-silenced plants showed a moderately reduced growth phenotype. Induction of hypersensitive cell death was compromised in the NbPDKs-silenced plants challenged with Ralstonia solanacearum. The hypersensitive cell death induced by bacterial effectors was also reduced in the NbPDKs-silenced plants. the NbPDKs-silenced plants showed decreased production of salicylic acid, jasmonic acid and jasmonoyl-L-isoleucine, as well as hydrogen peroxide after inoculation with R. solanacearum. These results suggest that NbPDKs might have an important role in the regulation of the hypersensitive cell death via plant hormone signaling and oxidative burst.
Camelina sativa is a Brassicaceae oilseed plant used as a biotechnology platform for biofuel and healthy vegetable oil. As Camelina is closely related to the model plant Arabidopsis, the genetic tools of Arabidopsis are considered useful when applied to Camelina. Myosin XI-2 is one of the major motive forces driving cytoplasmic streaming in Arabidopsis. In our previous study, high-speed chimeric myosin XI-2, a myosin XI-2 artificially modified by genetically exchanging the motor domain of Arabidopsis myosin XI-2 with the faster Chara myosin XI, was shown to accelerate cytoplasmic streaming and promote plant growth in Arabidopsis. Here, we heterologously transformed this high-speed Chara-Arabidopsis chimeric myosin XI-2 gene in Camelina. The transgenic plants exhibited not only enhancement of leaf development and main stem elongation but also early flowering and seed setting, indicating that the high-speed chimeric myosin XI-2 can improve plant growth in Camelina. Interestingly, total seed yield was significantly increased in the transgenic plants as the total seed number increased. Our results suggest that the high-speed myosin XI system might also be effective to improve the growth of other closely related plant species.