Arabidopsis ADC1 functions as an N~δ-acetylornithine decarboxylase

Polyamines are small aliphatic amines found in almost all organisms, ranging from bacteria to plants and animals. In most plants, putrescine, the metabolic precursor for longer polyamines, such as spermidine and spermine, is produced from arginine, with either agmatine or ornithine as intermediates. Here we show that Arabidopsis thaliana(Arabidopsis) arginine decarboxylase 1(ADC1), one of the two known arginine decarboxylases in Arabidopsis, not only synthesizes agmatine from arginine, but also converts N~δ-acetylornithine to N-acetylputrescine. Phylogenetic analyses indicate that duplication and neofunctionalization of ADC1 and NATA1, the enzymes that synthesize N~δ-acetylornithine in Arabidopsis, co-occur in a small number of related species in the Brassicaceae. Unlike ADC2, which is localized in the chloroplasts, ADC1 is in the endoplasmic reticulum together with NATA1, an indication that these two enzymes have access to the same substrate pool. Together, these results are consistent with a model whereby NATA1 and ADC1 together provide a pathway for the synthesis of N-acetylputrescine in Arabidopsis.     

Tillering and small grain 1 dominates the tryptophan aminotransferase family required for local auxin biosynthesis in rice

Auxin is a crucial phytohormone, controlling multiple aspects of plant growth and responses to the changing environment. However, the role of local auxin biosynthesis in specific developmental programs remains unknown in crops. This study characterized the rice tillering and small grain 1 (tsg1) mutant, which has more tillers but a smaller panicle and grain size resulting from a reduction in endogenous auxin. TSG1 encodes a tryptophan aminotransferase that is allelic to the FISH BONE (FIB) gene. The tsg1 mutant showed hypersensitivity to indole‐3‐acetic acid and the competitive inhibitor of aminotransferase, L‐kynurenine. TSG1 knockout resulted in an increased tiller number but reduction in grain number and size, and decrease in height. Meanwhile, deletion of the TSG1 homologs OsTAR1, OsTARL1, and OsTARL2 caused no obvious changes, although the phenotype of the TSG1/OsTAR1 double mutant was intensified and infertile, suggesting gene redundancy in the rice tryptophan aminotransferase family. Interestingly, TSG1 and OsTAR1, but not OsTARL1 and OsTARL2, displayed marked aminotransferase activity. Meanwhile, subcellular localization was identified as the endoplasmic reticulum, while phylogenetic analysis revealed functional divergence of TSG1 and OsTAR1 from OsTARL1 and OsTARL2. These findings suggest that TSG1 dominates the tryptophan aminotransferase family, playing a prominent role in local auxin biosynthesis in rice.     

Cryptochrome‐mediated hypocotyl phototropism was regulated antagonistically by gibberellic acid and sucrose in Arabidopsis

Both phototropins(phot1 and phot2) and cryptochromes(cry1 and cry2) were proven as the Arabidopsis thaliana blue light receptors. Phototropins predominately function in photomovement, and cryptochromes play a role in photomorphogenesis. Although cryptochromes have been proposed to serve as positive modulators of phototropic responses, the underlying mechanism remains unknown. Here, we report that depleting sucrose from the medium or adding gibberellic acids(GAs) can partially restore the defects in phototropic curvature of the phot1 phot2 double mutants under high-intensity blue light; this restoration does not occur in phot1 phot2 cry1 cry2 quadruple mutants and nph3(nonphototropic hypocotyl 3) mutants which were impaired phototropic response in sucrose-containing medium. These results indicate that GAs and sucrose antagonistically regulate hypocotyl phototropism in a cryptochromes dependent manner, but it showed a crosstalk with phototropin signaling on NPH3.Furthermore, cryptochromes activation by blue light inhibit GAs synthesis, thus stabilizing DELLAs to block hypocotyl growth, which result in the higher GAs content in the shade side than the lit side of hypocotyl to support the asymmetric growth of hypocotyl. Through modulation of the abundance of DELLAs by sucrose depletion or added GAs, it revealed that cryptochromes have a function in mediating phototropic curvature.     

Natural variation in the promoter of OsHMA3 contributes to differential grain cadmium accumulation between Indica and Japonica rice

Rice is a major source of cadmium(Cd) intake for Asian people. Indica rice usually accumulates more Cd in shoots and grains than Japonica rice. However, underlying genetic bases for differential Cd accumulation between Indica and Japonica rice are still unknown. In this study, we cloned a quantitative trait locus(QTL) grain Cd concentration on chromosome 7(GCC7) responsible for differential grain Cd accumulation between two rice varieties by performing QTL analysis and map-based cloning. We found that the two GCC7 alleles, GCC7~(PA64s) and GCC7~(93-11), had different promoter activity of OsHMA3,leading to different OsHMA3 expression and different shoot and grain Cd concentrations. By analyzing the distribution of different haplotypes of GCC7 among diverse rice accessions, we discovered that the high and low Cd accumulation alleles, namely GCC7~(93-11) and GCC7~(PA64s), were preferentially distributed in Indica and Japonica rice,respectively. We further showed that the GCC7~(PA64s)allele can be used to replace the GCC7~(93-11) allele in the super cultivar 93-11 to reduce grain Cd concentration without adverse effect on agronomic traits. Our results thus reveal that the QTL GCC7 with sequence variation in the OsHMA3 promoter is an important determinant controlling differential grain Cd accumulation between Indica and Japonica rice.     

Split Nano luciferase complementation for probing protein‐protein interactions in plant cells

Deciphering protein‐protein interactions (PPIs) is fundamental for understanding signal transduction pathways in plants. The split firefly luciferase (Fluc) complementation (SLC) assay has been widely used for analyzing PPIs. However, concern has risen about the bulky halves of Fluc interfering with the functions of their fusion partners. Nano luciferase (Nluc) is the smallest substitute for Fluc with improved stability and luminescence. Here, we developed a dual‐use system enabling the detection of PPIs through the Nluc‐based SLC and co‐immunoprecipitation assays. This was realized by coexpression of two proteins under investigation in fusion with the HA‐ or FLAG‐tagged Nluc halves, respectively. We validated the robustness of this system by reproducing multiple previously documented PPIs in protoplasts or Agrobacterium‐transformed plants. We next applied this system to evaluate the homodimerization of Arabidopsis CERK1, a coreceptor of fungal elicitor chitin, and its heterodimerization with other homologs in the absence or presence of chitin. Moreover, split fragments of Nluc were fused to two cytosolic ends of Arabidopsis calcium channels CNGC2 and CNGC4 to help sense the allosteric change induced by the bacterial elicitor flg22. Collectively, these results demonstrate the usefulness of the Nluc‐based SLC assay for probing constitutive or inducible PPIs and protein allostery in plant cells.     

A combined CaMKII inhibition and mineralocorticoid receptor antagonism via eplerenone inhibits functional deterioration in chronic pressure overloaded mice

In the diseased and remodelled heart, increased activity and expression of Ca^2+/calmodulin‐dependent protein kinase II (CaMKII), an excess of fibrosis, and a decreased electrical coupling and cellular excitability leads to disturbed calcium homeostasis and tissue integrity. This subsequently leads to increased arrhythmia vulnerability and contractile dysfunction. Here, we investigated the combination of CaMKII inhibition (using genetically modified mice expressing the autocamtide‐3‐related‐peptide (AC3I)) together with eplerenone treatment (AC3I‐Epler) to prevent electrophysiological remodelling, fibrosis and subsequent functional deterioration in a mouse model of chronic pressure overload. We compared AC3I‐Epler mice with mice only subjected to mineralocorticoid receptor (MR) antagonism (WT‐Epler) and mice with only CaMKII inhibition (AC3I‐No). Our data show that a combined CaMKII inhibition together with MR antagonism mitigates contractile deterioration as was manifested by a preservation of ejection fraction, fractional shortening, global longitudinal strain, peak strain and contractile synchronicity. Furthermore, patchy fibrosis formation was reduced, potentially via inhibition of pro‐fibrotic TGF‐β/SMAD3 signalling, which related to a better global contractile performance and a slightly depressed incidence of arrhythmias. Furthermore, the level of patchy fibrosis appeared significantly correlated to eplerenone dose. The addition of eplerenone to CaMKII inhibition potentiates the effects of CaMKII inhibition on pro‐fibrotic pathways. As a result of the applied strategy, limiting patchy fibrosis adheres to a higher synchronicity of contraction and an overall better contractile performance which fits with a tempered arrhythmogenesis.