The rice CYP78A gene BSR2 confers resistance to Rhizoctonia solani and affects seed size and growth in Arabidopsis and rice.

The fungal pathogen Rhizoctonia solani causes devastating diseases in hundreds of plant species. Among these, R. solani causes sheath blight, one of the three major diseases in rice. To date, few genes have been reported that confer resistance to R. solani. Here, rice-FOX Arabidopsis lines identified as having resistance to a bacterial pathogen, Pseudomonas syringae pv. tomato DC3000, and a fungal pathogen, Colletotrichum higginsianum were screened for disease resistance to R. solani. BROAD-SPECTRUM RESISTANCE2 (BSR2), a gene encoding an uncharacterized cytochrome P450 protein belonging to the CYP78A family, conferred resistance to R. solani in Arabidopsis. When overexpressed in rice, BSR2 also conferred resistance to two R. solani anastomosis groups. Both Arabidopsis and rice plants overexpressing BSR2 had slower growth and produced longer seeds than wild-type control plants. In contrast, BSR2-knockdown rice plants were more susceptible to R. solani and displayed faster growth and shorter seeds in comparison with the control. These results indicate that BSR2 is associated with disease resistance, growth rate and seed size in rice and suggest that its function is evolutionarily conserved in both monocot rice and dicot Arabidopsis.

Rapid profiling method for mammalian feces short chain fatty acids by GC-MS.

Short chain fatty acids (SCFAs) are key feces metabolites generated by gut bacteria fermentation. Despite the importance of profiling feces SCFAs, technical difficulties in analysis remain due to their volatility and hydrophilicity. We improve previous protocols to profile SCFAs and optimize the metabolite profiling platform for mammalian feces samples. In this study, we investigated feces as biological samples using gas chromatography-mass spectrometry (GC-MS). Isobutyl chloroformate was used for a derivatization in aqueous solution without drying out the samples. Ultimately, we envisage being able to determine the way in which gut bacteria fermentation influences host gut condition by using our rapid metabolite profiling methods.

The reduction in maize leaf growth under mild drought affects the transition between cell division and cell expansion and cannot be restored by elevated gibberellic acid levels.

Growth is characterized by the interplay between cell division and cell expansion, two processes that occur separated along the growth zone at the maize leaf. To gain further insight into the transition between cell division and cell expansion, conditions were investigated in which the position of this transition zone was positively or negatively affected. High levels of gibberellic acid (GA) in plants overexpressing the GA biosynthesis gene GA20-OXIDASE (GA20OX-1OE ) shifted the transition zone more distally, whereas mild drought, which is associated with lowered GA biosynthesis, resulted in a more basal positioning. However, the increased levels of GA in the GA20OX-1OE line were insufficient to convey tolerance to the mild drought treatment, indicating that another mechanism in addition to lowered GA levels is restricting growth during drought. Transcriptome analysis with high spatial resolution indicated that mild drought specifically induces a reprogramming of transcriptional regulation in the division zone. 'Leaf Growth Viewer' was developed as an online searchable tool containing the high-resolution data.

Abnormal Endogenous Repression of GA Signaling in a Seedless Table Grape Cultivar with High Berry Growth Response to GA Application.

Gibberellin (GA) application is routinely used in the table grape industry to increase berry size and cluster length. Although grapevine cultivars show a wide range of growth responsiveness to GA3 application, the reasons for these differences is unclear. To shed light on this issue, two commercial grapevine cultivars with contrasting berry response to GA were selected for comparative analysis, in which we tested if the differences in response: (1) is organ-specific or cultivar-related; (2) will be reflected in qualitative/quantitative differences in transcripts/proteins of central components of GA metabolism and signaling and levels of GA metabolites. Our results showed that in addition to the high response of its berries to GA, internodes and rachis of cv. Black finger (BF) presented a greater growth response compared to that of cv. Spring blush (SB). In agreement, the results exposed significant quantitative differences in GA signaling components in several organs of both cultivars. Exceptionally higher level of all three functional VvDELLA proteins was recorded in young BF organs, accompanied by elevated VvGID1 expression and lower VvSLY1b transcripts. Absence of seed traces, low endogenous GA quantities and lower expression of VvGA20ox4 and VvGA3ox3 were also recorded in berries of BF. Our results raise the hypothesis that, in young organs of BF, low expression of VvSLY1b may be responsible for the massive accumulation of VvDELLA proteins, which then leads to elevated VvGID1 levels. This integrated analysis suggests causal relationship between endogenous mechanisms leading to anomalous GA signaling repression in BF, manifested by high quantities of VvDELLA proteins, and greater growth response to GA application.

Analysis of plant hormone profiles in response to moderate dehydration stress.

Plant responses to dehydration stress are mediated by highly complex molecular systems involving hormone signaling and metabolism, particularly the major stress hormone abscisic acid (ABA) and ABA-dependent gene expression. To understand the roles of plant hormones and their interactions during dehydration, we analyzed the plant hormone profiles with respect to dehydration responses in Arabidopsis thaliana wild-type (WT) plants and ABA biosynthesis mutants (nced3-2). We developed a procedure for moderate dehydration stress, and then investigated temporal changes in the profiles of ABA, jasmonic acid isoleucine (JA-Ile), salicylic acid (SA), cytokinin (trans-zeatin, tZ), auxin (indole-acetic acid, IAA), and gibberellin (GA4 ), along with temporal changes in the expression of key genes involved in hormone biosynthesis. ABA levels increased in a bi-phasic pattern (at the early and late phases) in response to moderate dehydration stress. JA-Ile levels increased slightly in WT plants and strongly increased in nced3-2 mutant plants at 72 h after the onset of dehydration. The expression profiles of dehydration-inducible genes displayed temporal responses in an ABA-dependent manner. The early phase of ABA accumulation correlated with the expression of touch-inducible genes and was independent of factors involved in the major ABA regulatory pathway, including the ABA-responsive element-binding (AREB/ABF) transcription factor. JA-Ile, SA, and tZ were negatively regulated during the late dehydration response phase. Transcriptome analysis revealed important roles for hormone-related genes in metabolism and signaling during dehydration-induced plant responses.

Highly Sprouting-Tolerant Wheat Grain Exhibits Extreme Dormancy and Cold Imbibition-Resistant Accumulation of Abscisic Acid.

Pre-harvest sprouting (PHS) of wheat (Triticum aestivum L.) grains induces hydrolyzing enzymes such as α-amylase, which considerably decreases wheat product quality. PHS occurs when cool and wet weather conditions before harvest break dormancy and induce grain germination. In this study, we used PHS-tolerant varieties, Gifu-komugi (Gifu) and OS38, to characterize the mechanisms of both dormancy breakage and dormancy maintenance at low temperatures. Physiologically mature Gifu grains exhibited dormancy after imbibition at 20°C, but germinated at 15°C. In contrast, OS38 grains remained dormant even at temperatures as low as 5°C. Embryo half-grains cut out from the dormant Gifu grains germinated by imbibition at 20°C, similar to conventional varieties worldwide. However, OS38 embryo half-grains were still dormant. Hormonome and pharmacological analyses suggested that ABA and gibberellin metabolism are important for temperature-dependent dormancy maintenance and breakage. Imbibition at 15°C decreased ABA levels but increased gibberellin levels in embryos of freshly harvested Gifu grains. Additionally, low temperatures induced expression of the ABA catabolism genes,TaABA8' OH1 and TaABA8' OH2, and the gibberellin biosynthesis gene,TaGA3ox2, in the embryos. However, in embryos of freshly harvested OS38 grains, ABA levels were increased while gibberellin levels were suppressed at 15°C. In these dormant embryos, low temperatures induced the TaNCED ABA biosynthesis genes, but suppressed TaABA8' OH2 and TaGA3ox2.These results show that the regulatory mechanism influencing the expression of ABA and gibberellin metabolism genes may be critical for dormancy maintenance and breakage at low temperatures. Our findings should help improve PHS-resistant wheat breeding programs.

A chloroplast-localized protein LESION AND LAMINA BENDING affects defence and growth responses in rice.

Understanding how plants allocate their resources to growth or defence is of long-term importance to the development of new and improved varieties of different crops. Using molecular genetics, plant physiology, hormone analysis and Next-Generation Sequencing (NGS)-based transcript profiling, we have isolated and characterized the rice (Oryza sativa) LESION AND LAMINA BENDING (LLB) gene that encodes a chloroplast-targeted putative leucine carboxyl methyltransferase. Loss of LLB function results in reduced growth and yield, hypersensitive response (HR)-like lesions, accumulation of the antimicrobial compounds momilactones and phytocassanes, and constitutive expression of pathogenesis-related genes. Consistent with these defence-associated responses, llb shows enhanced resistance to rice blast (Magnaporthe oryzae) and bacterial blight (Xanthomonas oryzae pv. oryzae). The lesion and resistance phenotypes are likely to be caused by the over-accumulation of jasmonates (JAs) in the llb mutant including the JA precursor 12-oxo-phytodienoic acid. Additionally, llb shows an increased lamina inclination and enhanced early seedling growth due to elevated brassinosteroid (BR) synthesis and/or signalling. These findings show that LLB functions in the chloroplast to either directly or indirectly repress both JA- and BR-mediated responses, revealing a possible mechanism for controlling how plants allocate resources for defence and growth.

Enhancement of hypocotyl elongation by LOV KELCH PROTEIN2 production is mediated by auxin and phytochrome-interacting factors in Arabidopsis thaliana.

KEY MESSAGE: Auxin and two phytochrome-interacting factors, PHYTOCHROME-INTERACTING FACTOR4 (PIF4) and PIF5, play crucial roles in the enhancement of hypocotyl elongation in transgenic Arabidopsis thaliana plants that overproduce LOV KELCH PROTEIN2 (LKP2). LOV KELCH PROTEIN2 (LKP2) is a positive regulator of hypocotyl elongation under white light in Arabidopsis thaliana. In this study, using microarray analysis, we compared the gene expression profiles of hypocotyls of wild-type Arabidopsis (Columbia accession), a transgenic line that produces green fluorescent protein (GFP), and two lines that produce GFP-tagged LKP2 (GFP-LKP2). We found that, in GFP-LKP2 hypocotyls, 775 genes were up-regulated, including 36 auxin-responsive genes, such as 27 SMALL AUXIN UP RNA (SAUR) and 6 AUXIN/INDOLE-3-ACETIC ACID (AUX/IAA) genes, and 21 genes involved in responses to red or far-red light, including PHYTOCHROME-INTERACTING FACTOR4 (PIF4) and PIF5; and 725 genes were down-regulated, including 15 flavonoid biosynthesis genes. Hypocotyls of GFP-LKP2 seedlings, but not cotyledons or roots, contained a higher level of indole-3-acetic acid (IAA) than those of control seedlings. Auxin inhibitors reduced the enhancement of hypocotyl elongation in GFP-LKP2 seedlings by inhibiting the increase in cortical cell number and elongation of the epidermal and cortical cells. The enhancement of hypocotyl elongation was completely suppressed in progeny of the crosses between GFP-LKP2 lines and dominant gain-of-function auxin-resistant mutants (axr2-1 and axr3-1) or loss-of-function mutants pif4, pif5, and pif4 pif5. Our results suggest that the enhancement of hypocotyl elongation in GFP-LKP2 seedlings is due to the elevated level of IAA and to the up-regulated expression of PIF4 and PIF5 in hypocotyls.

Overaccumulation of γ-Glutamylcysteine in a Jasmonate-Hypersensitive Arabidopsis Mutant Causes Jasmonate-Dependent Growth Inhibition.

Glutathione (GSH) is essential for many aspects of plant biology and is associated with jasmonate signaling in stress responses. We characterized an Arabidopsis (Arabidopsis thaliana) jasmonate-hypersensitive mutant (jah2) with seedling root growth 100-fold more sensitive to inhibition by the hormone jasmonyl-isoleucine than the wild type. Genetic mapping and genome sequencing determined that the mutation is in intron 6 of GLUTATHIONE SYNTHETASE2, encoding the enzyme that converts γ-glutamylcysteine (γ-EC) to GSH. The level of GSH in jah2 was 71% of the wild type, while the phytoalexin-deficient2-1 (pad2-1) mutant, defective in GSH1 and having only 27% of wild-type GSH level, was not jasmonate hypersensitive. Growth defects for jah2, but not pad2, were also seen in plants grown to maturity. Surprisingly, all phenotypes in the jah2 pad2-1 double mutant were weaker than in jah2. Quantification of γ-EC indicated these defects result from hyperaccumulation of this GSH precursor by 294- and 65-fold in jah2 and the double mutant, respectively. γ-EC reportedly partially substitutes for loss of GSH, but growth inhibition seen here was likely not due to an excess of total glutathione plus γ-EC because their sum in jah2 pad2-1 was only 16% greater than in the wild type. Further, the jah2 phenotypes were lost in a jasmonic acid biosynthesis mutant background, indicating the effect of γ-EC is mediated through jasmonate signaling and not as a direct result of perturbed redox status.

The control of tomato fruit elongation orchestrated by sun, ovate and fs8.1 in a wild relative of tomato.

Within the cultivated tomato germplasm, sun, ovate and fs8.1 are the three predominant QTLs controlling fruit elongation. Although SUN and OVATE have been cloned, their role in plant growth and development are not well understood. To compare and contrast the effects of the three QTLs in a homogeneous background, we developed near isogenic lines (NILs) in the wild species Solanum pimpinellifolium LA1589 background. We carried out detailed morphological characterization of reproductive and vegetative organs in the single, double and triple NILs and determined the epistatic interactions of the three loci affecting fruit shape. The phenotypic evaluations demonstrated that the three loci regulate unique aspects of ovary and fruit elongation and in different temporal manners. The strongest effect on organ shape was caused by sun. In addition to fruit shape, sun also affected leaf and sepal elongation and stem thickness. The synergistic interaction between sun and ovate or fs8.1 suggested that the pathways involving SUN, OVATE and the gene(s) underlying fs8.1 may converge at a common node. The results of an extensive profiling analysis suggested that the degree of fruit elongation was not related to the accumulation of any of the classical hormones.