Sucrose Signaling Regulates Anthocyanin Biosynthesis Through a MAPK Cascade in Arabidopsis thaliana.

Anthocyanin accumulation specifically depends on sucrose (Suc) signaling. However, the molecular basis of this process remains unknown. In this study, in vitro pull-down assays identified ETHYLENE-INSENSITIVE3 (EIN3), a component of both sugar signaling or/and metabolism. This protein interacted with YDA, and the physiological relevance of this interaction was confirmed by in planta co-immunoprecipitation, yeast two-hybrid (Y2H) assay, and bimolecular fluorescence complementation. Ethylene insensitive3-like 1 (eil1) ein3 double-mutant seedlings, but not ein3-1 seedlings, showed anthocyanin accumulation. Furthermore, ein3-1 suppressed anthocyanin accumulation in yda-1 plants. Thus, EMB71/YDA-EIN3-EIL1 may form a sugar-mediated gene cascade integral to the regulation of anthocyanin accumulation. Moreover, the EMB71/YDA-EIN3-EIL1 gene cascade module directly targeted the promoter of Transparent Testa 8 (TT8) by direct EIN3 binding. Collectively, our data inferred a molecular model where the signaling cascade of the YDA-EIN3-TT8 appeared to target TT8 via EIN3, thereby modulating Suc signaling-mediated anthocyanin accumulation.

Purification and identification of an actinomycin D analogue from actinomycetes associated with Ganoderma applanatum via magnetic molecularly imprinted polymers and tandem mass spectrometry.

Actinomycetes are main producers of antibiotics and targeted screening could improve the efficiency of discovering new drugs. This study describes, for the first time, the isolation of endophytic actinomycetes from the macrofungus Ganoderma applanatum. To increase the efficiency of screening, novel actinomycin D (Act D) molecularly-imprinted polymers were adsorbed to the surface of Fe3O4@SiO2 magnetic microspheres (MMIPs) and using in the isolation. A monolithic column prepared with magnetic molecularly imprinted polymers was employed to adsorb actinomycin D and its analogues for selective analysis and identification via MS/MS spectroscopy. The MMIP-monolithic column was selective for the structural features of Act D and its analogue, and the maximum loading of the MMIPs for Act D was ∼23.5 µg/g. The recognition time of the Act D was 20-30 min and had good discriminative ability. A new analogue was identified from endophytic actinomycetes KLBMP 2541, and it was purified using MMIPs comparison with MMIPs-solid phase extraction. Structural identification analysis confirmed that the new analogue was 2-methyl-actinomycin D, which has better anti-tumor activity than Act D. The presented method combines the advantages of MMIPs and MS with popular solutions to enable high affinity and selectivity screening of specific antibiotics from endophytic actinomycetes.

Arabidopsis ANGUSTIFOLIA3 (AN3) is associated with the promoter of CONSTITUTIVE PHOTOMORPHOGENIC1 (COP1) to regulate light-mediated stomatal development.

Light signals are perceived by multiple photoreceptors that converge to suppress the RING E3 ubiquitin ligase CONSTITUTIVE PHOTOMORPHOGENIC1 (COP1) for the regulation of stomatal development. Thus, COP1 is a point of integration between light signaling and stomatal patterning. However, how light signaling is collected into COP1 for the production and spacing of stomata is still unknown. Here, we report that the loss-of-function mutant of ANGUSTIFOLIA3 (AN3) delays asymmetric cell division, which leads to decreased stomatal index. Furthermore, overexpression of AN3 accelerates asymmetric cell division, which results in clusters of stomata. In addition, the stomatal development through AN3 regulation is mediated by light signaling. Finally, we find that an3 is a light-signaling mutant, and that AN3 protein is light regulated. Self-activation by AN3 contributes to the control of AN3 expression. Thus, AN3 is a point of collection between light signaling and stomatal patterning. Target-gene analysis indicates that AN3 is associated with COP1 promoter for the regulation of light-controlling stomatal development. Together, these components for regulating stomatal development form an AN3-COP1-E3 ubiquitin ligase complex, allowing the integration of light signaling into the production and spacing of stomata.

Integration of Environmental and Developmental (or Metabolic) Control of Seed Mass by Sugar and Ethylene Metabolisms in Arabidopsis.

In higher plants, seed mass is an important to evolutionary fitness. In this context, seedling establishment positively correlates with seed mass under conditions of environmental stress. Thus, seed mass constitutes an important agricultural trait. Here, we show loss-of-function of YODA (YDA), a MAPKK Kinase, and decreased seed mass, which leads to susceptibility to drought. Furthermore, we demonstrate that yda disrupts sugar metabolisms but not the gaseous plant hormone, ethylene. Our data suggest that the transcription factor EIN3 (ETHYLENE-INSENSITIVE3), integral to both sugar and ethylene metabolisms, physically interacts with YDA. Further, ein3-1 mutants exhibited increased seed mass. Genetic analysis indicated that YDA and EIN3 were integral to a sugar-mediated metabolism cascade which regulates seed mass by maternally controlling embryo size. It is well established that ethylene metabolism leads to the suppression of drought tolerance by the EIN3 mediated inhibition of CBF1, a transcription factor required for the expression genes of abiotic stress. Our findings help guide the synthesis of a model predicting how sugar/ethylene metabolisms and environmental stress are integrated at EIN3 to control both the establishment of drought tolerance and the production of seed mass. Collectively, these insights into the molecular mechanism underpinning the regulation of plant seed size may aid prospective breeding or design strategies to increase crop yield.

Soluble Sugar Accumulation Can Influence Seed Size via AN3-YDA Gene Cascade.

In higher plants, seed size is central to many aspects in evolutionary fitness and is a crucial agricultural trait. In this study, Arabidopsis an3 (angustifolia3) mutants present with increased seed size. Target-gene analysis revealed that YDA, which encodes a mitogen-activated protein kinase kinase kinase, is a target gene of AN3. Indeed, the loss of YDA function decreases seed size. Furthermore, AN3 and YDA mutations both disrupt normal sucrose and glucose contents and cause altered seed size in an3 or yda mutants. With these results, we provide a molecular model in which soluble sugar accumulation might affect seed size regulation via the AN3-YDA gene cascade. Our findings guide the synthesis of a model that predicts the integration of soluble sugar accumulation at AN3 to control the establishment of seed size.