RPG1-B-Derived Resistance to AvrB-Expressing Pseudomonas syringae Requires RIN4-Like Proteins in Soybean

Soybean (Glycine max) RPG1-B (for resistance to Pseudomonas syringae pv glycinea) mediates species-specific resistance to P. syringae expressing the avirulence protein AvrB, similar to the nonorthologous RPM1 in Arabidopsis (Arabidopsis thaliana). RPM1-derived signaling is presumably induced upon AvrB-derived modification of the RPM1-interacting protein, RIN4 (for RPM1-interacting 4). We show that, similar to RPM1, RPG1-B does not directly interact with AvrB but associates with RIN4-like proteins from soybean. Unlike Arabidopsis, soybean contains at least four RIN4-like proteins (GmRIN4a to GmRIN4d). GmRIN4b, but not GmRIN4a, complements the Arabidopsis rin4 mutation. Both GmRIN4a and GmRIN4b bind AvrB, but only GmRIN4b binds RPG1-B. Silencing either GmRIN4a or GmRIN4b abrogates RPG1-B-derived resistance to P. syringae expressing AvrB. Binding studies show that GmRIN4b interacts with GmRIN4a as well as with two other AvrB/RPG1-B-interacting isoforms, GmRIN4c and GmRIN4d. The lack of functional redundancy among GmRIN4a and GmRIN4b and their abilities to interact with each other suggest that the two proteins might function as a heteromeric complex in mediating RPG1-B-derived resistance. Silencing GmRIN4a or GmRIN4b in rpg1-b plants enhances basal resistance to virulent strains of P. syringae and the oomycete Phytophthora sojae. Interestingly, GmRIN4a- or GmRIN4b-silenced rpg1-b plants respond differently to AvrB-expressing bacteria. Although both GmRIN4a and GmRIN4b function to monitor AvrB in the presence of RPG1-B, GmRIN4a, but not GmRIN4b, negatively regulates AvrB virulence activity in the absence of RPG1-B.One of the myriad plant defense responses activated upon pathogen invasion is signaling induced via the activation of resistance (R) proteins. R gene-mediated resistance is generally activated in response to race-specific pathogen effectors, termed avirulence proteins (Avr), and often results in the development of a hypersensitive reaction at the site of pathogen entry (Dangl et al., 1996). The hypersensitive reaction is a form of programmed cell death that results in the formation of necrotic lesions around the site of pathogen entry and is thought to help prevent pathogen spread by confining it to the dead cells.A majority of the known R proteins contain conserved structural domains, including N-terminal coiled coil (CC) or Toll-interleukin 1 receptor (TIR)-like domains, central nucleotide-binding site (NBS), and C-terminal Leu-rich repeat (LRR) domains (Martin et al., 2003). While some R proteins “perceive” pathogen presence via direct physical interactions with the cognate Avr proteins (Scofield et al., 1996; Jia et al., 2000; Leister and Katagiri, 2000; Deslandes et al., 2003), several others likely do so indirectly. This led to the suggestion that R proteins monitor the presence of Avr proteins by “guarding” other host proteins targeted by the pathogen effector (Van der Biezen and Jones, 1998; Innes, 2004; Jones and Dangl, 2006). Avr proteins enhance pathogen virulence in genetic backgrounds lacking cognate R proteins by targeting components of the host basal defense machinery, including “guardee” proteins (Chang et al., 2000; Guttman and Greenberg, 2001; Chen et al., 2004, Kim et al., 2005b; Ong and Innes, 2006; van Esse et al., 2007; Shan et al., 2008; Xiang et al., 2008). However, some Avr proteins were found to also target host proteins that do not contribute to the virulence function of the effector (Shang et al., 2006; Shabab et al., 2008; Zhou and Chai, 2008; Zipfel and Rathjen, 2008). This led to the proposition that plants express “decoy” proteins that mimic Avr-guardee recognition in the presence of the R protein. This decoy model suggests that, unlike guardees, decoy proteins do not directly contribute to host basal immunity, such that Avr-derived alterations of decoys do not enhance pathogen virulence in plants lacking the R protein (van der Hoorn and Kamoun, 2008).A well-studied example of an indirect mode of effector recognition is that of the Arabidopsis (Arabidopsis thaliana) R protein, RPM1 (for resistance to Pseudomonas syringae pv maculicola 1). RPM1 mediates resistance against bacteria expressing two different Avr proteins, AvrRpm1 (AvrRpm1_PmaM6) and AvrB (AvrB1_Pgyrace4). Although RPM1 does not directly interact with either AvrRpm1 or AvrB, it does associate with RIN4 (for RPM1-interacting 4), which interacts with AvrRpm1 and AvrB. RIN4 is required for RPM1-induced resistance to AvrRpm1/AvrB-expressing P. syringae (Mackey et al., 2002). Both AvrRpm1 and AvrB induce the phosphorylation of RIN4, which is thought to induce RPM1-mediated resistance signaling. RIN4 also associates with a second Arabidopsis R protein, RPS2 (for resistance to P. syringae), which mediates resistance against P. syringae expressing AvrRpt2. RPS2-mediated signaling is activated when AvrRpt2 (AvrRpt2_PtoJL1065), a Cys protease, cleaves RIN4 (Axtell and Staskawicz, 2003; Mackey et al., 2003; Kim et al., 2005a). The AvrRpt2-triggered loss of RIN4 compromises RPM1-mediated resistance, because RIN4 is not available for phosphorylation (Ritter and Dangl, 1996; Axtell and Staskawicz, 2003; Mackey et al., 2003).The avirulence effector AvrB was first isolated from a P. syringae strain colonizing soybean (Glycine max) and used to identify the cognate resistance locus RPG1 in soybean (Staskawicz et al., 1987; Keen and Buzzell, 1991). This locus contains the RPG1-B (for resistance to P. syringae pv glycinea) gene, which encodes a CC-NBS-LRR protein conferring resistance to AvrB-expressing P. syringae in soybean (Bisgrove et al., 1994; Ashfield et al., 2004). Unlike RPM1, RPG1-B does not confer specificity to AvrRpm1 (Ashfield et al., 1995). However, as in Arabidopsis, the soybean RPG1-B-derived hypersensitive reaction to AvrB-expressing bacteria is inhibited by the presence of AvrRpt2-expressing bacteria (Axtell and Staskawicz, 2003, Mackey et al., 2003; Ashfield et al., 2004). This suggests that RPG1-B and RPM1 might utilize common signaling components even though they share very limited sequence identity. Therefore, we investigated the possible involvement of RIN4-like proteins in RPG1-B-mediated resistance signaling. In addition to Arabidopsis, RIN4-like proteins have also been identified in tomato (Solanum lycopersicum) and lettuce (Lactuca sativa; Jeuken et al., 2009; Luo et al., 2009). In tomato, the NBS-LRR protein, Prf (for Pseudomonas resistance and fenthion sensitivity), and its interacting protein kinase, Pto, mediate resistance to the AvrPto (AvrPto1_PtoJL1065)-expressing strain of P. syringae (Scofield et al., 1996; Tang et al., 1996; Kim et al., 2002; Mucyn et al., 2006). AvrPto binds RIN4 proteins from both Arabidopsis (AtRIN4) and tomato (SlRIN4). Similar to AvrRpt2, AvrPto induces the proteolysis of RIN4, albeit only in the presence of Pto and Prf (Luo et al., 2009). However, in the case of AvrPto, degradation of RIN4 is the result of induced proteolytic activity in the plant, rather than that of AvrPto itself. In Lactuca (lettuce) species, the L. saligna RIN4 allele was recently shown to be essential for resistance to an avirulent strain of the downy mildew pathogen, Bremia lactucae (Jeuken et al., 2009).Here, we report that two functionally nonredundant isoforms of soybean RIN4 (GmRIN4) function in RPG1-B-derived resistance as well as in the virulence activity of AvrB in the absence of RPG1-B.     

Drought acclimation reduces O2*- accumulation and lipid peroxidation in wheat seedlings

Abiotic stresses cause ROS accumulation, which is detrimental to plant growth. It is well known that acclimation of plants under mild or sub-lethal stress condition leads to development of resistance in plants to severe or lethal stress condition. The generation of ROS and subsequent oxidative damage during drought stress is well documented in the crop plants. However, the effect of drought acclimation treatment on ROS accumulation and lipid peroxidation has not been examined so far. In this study, the effect of water stress acclimation treatment on superoxide radical (O(2)(-z.rad;)) accumulation and membrane lipid peroxidation was studied in leaves and roots of wheat (Triticum aestivum) cv. C306. EPR quantification of superoxide radicals revealed that drought acclimation treatment led to 2-fold increase in superoxide radical accumulation in leaf and roots with no apparent membrane damage. However under subsequent severe water stress condition, the leaf and roots of non-acclimated plants accumulated significantly higher amount of superoxide radicals and showed higher membrane damage than that of acclimated plants. Thus, acclimation-induced restriction of superoxide radical accumulation is one of the cellular processes that confers enhanced water stress tolerance to the acclimated wheat seedlings.     

Detoxification of Multiple Heavy Metals by a Half-Molecule ABC Transporter,HMT-1, and Coelomocytes of Caenorhabditis elegans

Background

Developing methods for protecting organisms in metal-polluted environments is contingent upon our understanding of cellular detoxification mechanisms. In this regard, half-molecule ATP-binding cassette (ABC) transporters of the HMT-1 subfamily are required for cadmium (Cd) detoxification. HMTs have conserved structural architecture that distinguishes them from other ABC transporters and allows the identification of homologs in genomes of different species including humans. We recently discovered that HMT-1 from the simple, unicellular organism, Schizosaccharomyces pombe, SpHMT1, acts independently of phytochelatin synthase (PCS) and detoxifies Cd, but not other heavy metals. Whether HMTs from multicellular organisms confer tolerance only to Cd or also to other heavy metals is not known.

Methodology/Principal Findings

Using molecular genetics approaches and functional in vivo assays we showed that HMT-1 from a multicellular organism, Caenorhabditis elegans, functions distinctly from its S. pombe counterpart in that in addition to Cd it confers tolerance to arsenic (As) and copper (Cu) while acting independently of pcs-1. Further investigation of hmt-1 and pcs-1 revealed that these genes are expressed in different cell types, supporting the notion that hmt-1 and pcs-1 operate in distinct detoxification pathways. Interestingly, pcs-1 and hmt-1 are co-expressed in highly endocytic C. elegans cells with unknown function, the coelomocytes. By analyzing heavy metal and oxidative stress sensitivities of the coelomocyte-deficient C. elegans strain we discovered that coelomocytes are essential mainly for detoxification of heavy metals, but not of oxidative stress, a by-product of heavy metal toxicity.

Conclusions/Significance

We established that HMT-1 from the multicellular organism confers tolerance to multiple heavy metals and is expressed in liver-like cells, the coelomocytes, as well as head neurons and intestinal cells, which are cell types that are affected by heavy metal poisoning in humans. We also showed that coelomocytes are involved in detoxification of heavy metals. Therefore, the HMT-1-dependent detoxification pathway and coelomocytes of C. elegans emerge as novel models for studies of heavy metal-promoted diseases.     

Effect of Plumbago zeylanica root powder induced preimplantationary loss and abortion on uterine luminal proteins in albino rats

P. zeylanica treatment during first 7 days of pregnancy abolished uterine proteins of 13,000, 19,000 and 26,000 and 75,000 Da molecular weights resulting in preimplantationary loss. Proteins having molecular weights 55,000 and 65,000 Da were absent in aborted rats, that were given P. zeylanica root powder since day 6 to day 17 of pregnancy. The results suggest that proteins having molecular weights 13,000, 19,000, 26,000 and 75,000 Da influence the implantation and proteins of 55,000 and 65,000 Da are required for the maintenance of the pregnancy.     

Antioxidant response of wheat roots to drought acclimation

Wheat (Triticum aestivum L.) seedlings of a drought-resistant cv. C306 were subjected to severe water deficit directly or through stress cycles of increasing intensity with intermittent recovery periods. The antioxidant defense in terms of redox metabolites and enzymes in root cells and mitochondria was examined in relation to membrane damage. Acclimated seedlings exhibited higher relative water content and were able to limit the accumulation of H₂O₂ and membrane damage during subsequent severe water stress conditions. This was due to systematic up-regulation of superoxide dismutase, ascorbate peroxidase (APX), catalase, peroxidases, and ascorbate–glutathione cycle components at both the whole cell level as well as in mitochondria. In contrast, direct exposure of severe water stress to non-acclimated seedlings caused greater water loss, excessive accumulation of H₂O₂ followed by elevated lipid peroxidation due to the poor antioxidant enzyme response particularly of APX, monodehydroascorbate reductase, dehydroascorbate reductase, glutathione reductase, and ascorbate–glutathione redox balance. Mitochondrial antioxidant defense was found to be better than the cellular defense in non-acclimated roots. Termination of stress followed by rewatering leads to a rapid enhancement in all the antioxidant defense components in non-acclimated roots, which suggested that the excess levels of H₂O₂ during severe water stress conditions might have inhibited or down-regulated the antioxidant enzymes. Hence, drought acclimation conferred enhanced tolerance toward oxidative stress in the root tissue of wheat seedlings due to both reactive oxygen species restriction and well-coordinated induction of antioxidant defense.     

The N-terminal extension domain of the C. elegans half-molecule ABC transporter, HMT-1, is required for protein-protein interactions and function

Background

Members of the HMT-1 (heavy metal tolerance factor 1) subfamily of the ATP-binding cassette (ABC) transporter superfamily detoxify heavy metals and have unique topology: they are half-molecule ABC transporters that, in addition to a single transmembrane domain (TMD1) and a single nucleotide-binding domain (NBD1), possess a hydrophobic NH2-terminal extension (NTE). These structural features distinguish HMTs from other ABC transporters in different species including Drosophila and humans. Functional ABC transporters, however, are comprised of at least four-domains (two TMDs and two NDBs) formed from either a single polypeptide or by the association of two or four separate subunits. Whether HMTs act as oligomers and what role the NTE domain plays in their function have not been determined.

Methodology/Principal Findings

In this study, we examined the oligomeric status of Caenorhabditis elegans HMT-1 and the functional significance of its NTE using gel-filtration chromatography in combination with the mating-based split-ubiquitin yeast two-hybrid system (mbSUS) and functional in vivo assays. We found that HMT-1 exists in a protein complex in C. elegans. Studies in S. cerevisiae showed that HMT-1 at a minimum homodimerizes and that oligomerization is essential for HMT-1 to confer cadmium tolerance. We also established that the NTE domain plays an important structural and functional role: it is essential for HMT-1 oligomerization and Cd-detoxification function. However, the NTE itself was not sufficient for oligomerization suggesting that multiple structural features of HMT-1 must associate to form a functional transporter.

Conclusions

The prominence of heavy metals as environmental toxins and the remarkable conservation of HMT-1 structural architecture and function in different species reinforce the value of continued studies of HMT-1 in model systems for identifying functional domains in HMT1 of humans.     

Glycerol-3-phosphate and systemic immunity

Glycerol-3-phosphate (G3P), a conserved three-carbon sugar, is an obligatory component of energy-producing reactions including glycolysis and glycerolipid biosynthesis. G3P can be derived via the glycerol kinase-mediated phosphorylation of glycerol or G3P dehydrogenase (G3Pdh)-mediated reduction of dihydroxyacetone phosphate. Previously, we showed G3P levels contribute to basal resistance against the hemibiotrophic pathogen, Colletotrichum higginsianum. Inoculation of Arabidopsis with C. higginsianum correlated with an increase in G3P levels and a concomitant decrease in glycerol levels in the host. Plants impaired in GLY1 encoded G3Pdh accumulated reduced levels of G3P after pathogen inoculation and showed enhanced susceptibility to C. higginsianum. Recently, we showed that G3P is also a potent inducer of systemic acquired resistance (SAR) in plants. SAR is initiated after a localized infection and confers whole-plant immunity to secondary infections. SAR involves generation of a signal at the site of primary infection, which travels throughout the plants and alerts the un-infected distal portions of the plant against secondary infections. Plants unable to synthesize G3P are defective in SAR and exogenous G3P complements this defect. Exogenous G3P also induces SAR in the absence of a primary pathogen. Radioactive tracer experiments show that a G3P derivative is translocated to distal tissues and this requires the lipid transfer protein, DIR1. Conversely, G3P is required for the translocation of DIR1 to distal tissues. Together, these observations suggest that the cooperative interaction of DIR1 and G3P mediates the induction of SAR in plants.Glycerol-3-phosphate (G3P) is an obligatory component of energy-producing reactions including glycolysis and glycerolipid biosynthesis.^1,2 G3P levels in the plant are regulated by enzymes directly/indirectly involved in G3P biosynthesis, as well as those involved in G3P catabolism. G3P is synthesized via the glycerol kinase (GK)-mediated phosphorylation of glycerol,³ or the G3P dehydrogenase (G3Pdh)-mediated reduction of dihydroxyacetone phosphate (DHAP)⁴ (Fig. 1). DHAP is derived from glycolysis via triosephosphate isomerase activity on glyceraldehyde-3-phosphate, or from the conversion of glycerol to dihydroxacetone (DHA) by glycerol dehydrogenase (Glydh) followed by phosphorylation of DHA to DHAP by DHA kinase (DHAK). G3P is catabolized either upon its conversion to glycerol by glycerol-3-phoshatase (GPP) or its utilization in glycerolipid/triacylglycerol biosynthesis. In Arabidopsis, the total G3P pool is derived from the activities of five G3Pdh isoforms and one GK isoform present in three cellular locations^5-9; GK and two of the G3Pdh isoforms are present in the cytoplasm, two other G3Pdh isoforms localize to plastids, and one to the mitochondria. One of the plastid localized G3Pdh isoforms, designated GLY1, was previously shown to be required for glycerolipid biosynthesis; a mutation in GLY1 compromised lipids synthesized via the plastidal pathway of lipid biosynthesis. The fact that exogenous application of glycerol to gly1 plants normalizes plastidal lipid levels¹⁰ and that GLY1 encodes a G3Pdh⁴ suggests that the G3P pool generated via the GLY1 catalyzed reaction is required for the biosynthesis of plastidal lipids. Intriguingly, unlike GLY1, neither the chloroplastic, nor the two cytosolic isoforms of G3Pdh, contribute to plastidal and/or extraplastidal lipid biosynthesis.⁹Open in a separate windowFigure 1.A condensed scheme of glycerol-3-phosphate metabolism in plants. Glycerol is phosphorylated to glycerol-3-phosphate (G3P) by glycerol kinase (GK; GLI1). G3P can also be generated by G3P dehydrogenase (G3Pdh) via the reduction of dihydroxyacetone phosphate (DHAP). DHAP is derived from glycolysis via triosephosphate isomerase (TPI) activity on glyceraldehyde-3-phosphate (Gld-3-P), or from the conversion of glycerol to dihydroxacetone (DHA) by glycerol dehydrogenase (Glydh) followed by phosphorylation of DHA to DHAP by DHA kinase (DHAK). G3Pdh isoforms are present in both the cytosol and the plastids (represented by the oval). GLY1 is one of the two plastidial G3Pdh isoforms that plays an important role in plastidial glycerolipid biosynthesis. In the plastids, G3P is acylated with oleic acid (18:1) by the ACT1-encoded G3P acyltransferase. This ACT1-utilized 18:1 is derived from the stearoyl-acyl carrier protein (ACP)-desaturase (SACPD)-catalyzed desaturation of stearic acid (18:0). The 18:1-ACP generated by SACPD either enters the prokaryotic lipid biosynthetic pathway through acylation of G3P or is exported out (dotted line) of the plastids as a coenzyme A (CoA)-thioester to enter the eukaryotic lipid biosynthetic pathway. Membranous fatty acid desaturases (FAD) catalyze desaturation of FAs present on membranous glycerolipids. Other abbreviations used are: GL, glycerolipid; FAS, fatty acid synthase; ACC, acetyl-CoA carboxylase; Lyso-PA, acyl-G3P; PA, phosphatidic acid; PG, phosphatidylglycerol; MGDG, monogalactosyldiacylglycerol; DGDG, digalactosyldiacylglycerol; SL, sulfolipid; DAG, diacylglycerol.For glycerolipid biosynthesis, G3P is first acylated with the fatty acid (FA) oleic acid (18:1), to form lyso-phosphatidic acid (lyso-PA) via the activity of the soluble G3P acyltransferase (GPAT) encoded by the ACT1 gene in Arabidopsis¹¹ (Fig. 1). 18:1 in turn is derived from the saturated FA, stearic acid (18:0), via the activity of soluble stearoyl-acyl carrier protein desaturases (SACPD),¹² which introduce a single cis double bond in 18:0. The 18:1 generated via this reaction is either exported out of the plastids or acylated at the sn-1 position of G3P. Previously, we have shown that 18:1 levels are important regulators of plant defense signaling. In Arabidopsis, 18:1 is synthesized via the SSI2/FAB2-encoded SACPD,¹² which uses 18:0 as a substrate. A mutation in SSI2 results in the accumulation of 18:0 and a reduction in 18:1 levels. The mutant plants show stunting, spontaneous lesion formation, constitutive PR gene expression, and enhanced resistance to bacterial and oomycete pathogens.^4,12-17 Characterization of ssi2 suppressor mutants has shown that the altered defense-related phenotypes are the result of the reduction in the levels of the unsaturated FA, 18:1, which causes induction of several resistance (R) genes.^4,14,18,19 Restoration of 18:1 levels, via mutations in ACT1,¹⁴ GLY1⁴ or ACP4,¹⁸ normalizes R gene expression in ssi2 plants. The low 18:1-mediated induction of R gene expression and the associated defense signaling can also be suppressed by simultaneous mutations in EDS1 and the genes governing salicylic acid (SA) biosynthesis (SID2, EDS5).¹⁹ Furthermore, the functional redundancy between EDS1 and SA likely masks the requirement for EDS1 by several coiled coil (CC)- nucleotide binding site (NBS)- leucine rich repeat (LRR) proteins,¹⁹ previously thought to function independent of EDS1.²⁰ Thus, the reliance on EDS1 for signaling mediated by CC-NBS-LRR proteins becomes evident only in the absence of SA.The plastidal 18:1 levels are also regulated via the chloroplastic G3P pool and vice-versa. However, 18:1 and G3P appear to function distinctly in defense signaling. For example, G3P levels are important for basal defense against the hemibiotrophic fungus, Colletotrichum higginsianum.^21,22 Genetic mutations affecting G3P synthesis in Arabidopsis enhance susceptibility to C. higginsianum. Conversely, plants accumulating increased G3P show enhanced resistance. More recently, we demonstrated roles for G3P in R-mediated defense leading to systemic acquired resistance (SAR).⁹ R-mediated defense against the avirulent bacterial pathogen P. syringae is associated with a rapid increase in G3P levels; G3P levels peak within 6 h of inoculation with avirulent bacteria (avrRpt2), in resistant plants expressing the R gene RPS2. Strikingly, accumulation of G3P, in the infected and systemic tissues, precedes the accumulation of other metabolites known to be essential for SAR; SA,^23,24 jasmonic acid (JA)²⁵ and azelaic acid (AA)²⁶ accumulated at least 24 h post pathogen inoculation. Furthermore, mutants defective in G3P synthesis are compromised in SAR but accumulated normal levels of SA, AA, and JA. Compromised SAR in G3P deficient mutants was restored by exogenous application of G3P, thus arguing a role for G3P in SAR. This was further supported by the fact that exogenous G3P induced SAR in the absence of the primary pathogen in both Arabidopsis and soybean.⁹ That fact that G3P is a conserved metabolite common to prokaryotes, plants, and humans further corroborates the conserved nature of SAR signaling. Interestingly, although exogenous G3P did not induce SA biosynthesis, SAR conferred by exogenous G3P was dependent on SA. These results suggest that the onset and/or establishment of SAR likely requires basal, but not induced levels of SA, in the distal tissues. It is possible that the relatively small increase in SA observed in the systemic tissues during SAR is an indirect response that contributes to generalized resistance, rather than SAR itself. Interestingly, both G3P conferred SAR, and the systemic movement of G3P were dependent on the lipid transfer protein, DIR1, a well-known positive regulator of SAR.²⁷ Conversely, systemic movement of DIR1 required G3P. These findings did not correlate with the fact that G3P is cytosolic while DIR1 was a predicted apoplastic protein. To resolve this issue, we studied the localization of DIR1, and found that it is in fact a symplastic protein. The symplastic location of DIR1 was further corroborated when GFP fused to the signal peptide from DIR1 localized to the endoplasmic reticulum, rather than the typical cytoplasmic and nuclear location of GFP (Fig. 2). These results suggested that the symplastic movement of DIR1 is likely critical for SAR, and supported the facts that G3P and DIR1 are interdependent for translocation to systemic tissues. However, these findings could not explain how a lipid transfer-like protein might associate with the phosphorylated sugar G3P, to move systemically. Analysis of G3P in the leaf extracts showed that it was derivatized into an unknown compound before/during translocation. It is likely that the G3P derivative has a lipid moiety via which it associates with DIR1 for transfer. In summary, we showed that DIR1 together with a G3P-derived compound are sufficient for the induction of SAR in wild type plants. Our findings provide strong evidence in support of a direct defense-signaling role for G3P and warranty further analysis of its metabolic pathway(s) for their role(s) in various modes of plant defense.Open in a separate windowFigure 2.Confocal micrograph showing localization of GFP fused to DIR1 transit peptide (TP) or GFP alone in Nicotiana benthamiana plants expressing RFP-tagged nuclear histone protein H2B. Arrow indicates nucleus, arrowhead indicates endoplasmic reticulum.     

A novel soft sensor approach for estimating individual biomass in mixed cultures

Due to many advantages associated with mixed cultures, their application in biotechnology has expanded rapidly in recent years. At the same time, many challenges remain for effective mixed culture applications. One obstacle is how to efficiently and accurately monitor the individual cell populations. Current approaches on individual cell mass quantification are suitable for off‐line, infrequent characterization. In this study, we propose a fast and accurate “soft sensor” approach for estimating individual cell concentrations in mixed cultures. The proposed approach utilizes optical density scanning spectrum of a mixed culture sample measured by a spectrophotometer over a range of wavelengths. A multivariate linear regression method, partial least squares or PLS, is applied to correlate individual cell concentrations to the spectrum. Three experimental case studies are used to examine the performance of the proposed soft sensor approach. © 2017 American Institute of Chemical Engineers Biotechnol. Prog., 33:347–354, 2017     

Drought-induced spikelet sterility is associated with an inefficient antioxidant defence in rice panicles

Water stress-induced spikelet sterility limits rice production under upland conditions. The causes of spikelet sterility under drought stress are poorly understood. In this study the role of antioxidant defence management in drought-induced spikelet sterility was investigated in two rice ( Oryza sativa ) genotypes differing in drought resistance. Drought-resistant N22 genotype showed less water stress-induced spikelet sterility when compared to the susceptible N118 genotype under upland conditions. The N22 panicles maintained higher RWC and turgor potential and lower H₂O₂ levels across the developmental stages under water stress than that of N118 panicles. Drought-induced enhancement in superoxide dismutase (SOD, EC 1.15.1.1) activity coupled with higher ascorbate (AsA), glutathione (GSH) content and enhanced ascorbate peroxidase (APX, EC 1.11.1.11) and glutathione reductase (GR, EC 1.6.4.2) activities resulted in lower H₂O₂ levels in N22 panicles. In contrast, insufficient enhancement in SOD, APX and GR activities resulted in relatively higher H₂O₂ levels under water stress in N118 panicles. The N22 panicles exhibited a higher number of SOD and APX isozymes in comparison with N118 panicles that might provide better reactive oxygen species scavenging. Hence it is concluded that well-equipped antioxidant defence plays an important role in minimizing water stress-induced spikelet sterility in upland rice.