Plant age-dependent dynamics of annatto pigment (bixin) biosynthesis in Bixa orellana.

Age affects the production of secondary metabolites, but how developmental cues regulate secondary metabolism remains poorly understood. The achiote tree (Bixa orellana L.) is a source of bixin, an apocarotenoid used in diverse industries worldwide. Understanding how age-dependent mechanisms control bixin biosynthesis is of great interest for plant biology and for economic reasons. Here we overexpressed miRNA156 (miR156) in B. orellana to comprehensively study the effects of the miR156-SQUAMOSA PROMOTER BINDING PROTEIN-LIKE (SPL) module on age-dependent bixin biosynthesis in leaves. Overexpression of miR156 in annatto plants (miR156ox) reduced BoSPL transcript levels, impacted leaf ontogeny, lessened bixin production, and increased abscisic acid levels. Modulation of expression of BoCCD4-4 and BoCCD1, key genes in carotenoid biosynthesis, was associated with diverting the carbon flux from bixin to abscisic acid in miR156ox leaves. Proteomic analyses revealed an overall low accumulation of most secondary metabolite-related enzymes in miR156ox leaves, suggesting that miR156-targeted BoSPLs may be required to activate several secondary metabolic pathways. Our findings suggest that the conserved BomiR156-BoSPL module is deployed to regulate leaf dynamics of bixin biosynthesis, and may create novel opportunities to fine-tune bixin output in B. orellana breeding programs.

Physiological, epigenetic, and proteomic responses in Pfaffia glomerata growth in vitro under salt stress and 5-azacytidine.

Plants adjust their complex molecular, biochemical, and metabolic processes to overcome salt stress. Here, we investigated the proteomic and epigenetic alterations involved in the morphophysiological responses of Pfaffia glomerata, a medicinal plant, to salt stress and the demethylating agent 5-azacytidine (5-azaC). Moreover, we investigated how these changes affected the biosynthesis of 20-hydroxyecdysone (20-E), a pharmacologically important specialized metabolite. Plants were cultivated in vitro for 40 days in Murashige and Skoog medium supplemented with NaCl (50 mM), 5-azaC (25 µM), and NaCl + 5-azaC. Compared with the control (medium only), the treatments reduced growth, photosynthetic rates, and photosynthetic pigment content, with increase in sucrose, total amino acids, and proline contents, but a reduction in starch and protein. Comparative proteomic analysis revealed 282 common differentially accumulated proteins involved in 87 metabolic pathways, most of them related to amino acid and carbohydrate metabolism, and specialized metabolism. 5-azaC and NaCl + 5-azaC lowered global DNA methylation levels and 20-E content, suggesting that 20-E biosynthesis may be regulated by epigenetic mechanisms. Moreover, downregulation of a key protein in jasmonate biosynthesis indicates the fundamental role of this hormone in the 20-E biosynthesis. Taken together, our results highlight possible regulatory proteins and epigenetic changes related to salt stress tolerance and 20-E biosynthesis in P. glomerata, paving the way for future studies of the mechanisms involved in this regulation.

Dark Septate Endophytic Fungi Increase Green Manure-15N Recovery Efficiency, N Contents, and Micronutrients in Rice Grains.

An understanding of the interaction between rice and dark septate endophytic (DSE) fungi, under green fertilization, may lead to sustainable agricultural practices. Nevertheless, this interaction is still poorly understood. Therefore, in this study, we aimed to evaluate the accumulation of macro- and micronutrients, dry matter, and protein and N recovery efficiency from Canavalia ensiformis (L.)-15N in rice inoculated with DSE fungi. An experiment under greenhouse conditions was conducted in a randomized complete block design comprising split-plots, with five replicates of rice plants potted in non-sterilized soil. Rice (Piauí variety) seedlings were inoculated with DSE fungi, A101 and A103, or left uninoculated (control) and transplanted into pots containing 12 kg of soil, which had previously been supplemented with dry, finely ground shoot biomass of C. ensiformis enriched with 2.15 atom % 15N. Two collections were performed in the experiment: one at 54 days after transplanting (DAT) and one at 130 DAT (at maturation). Growth indicators (at 54 DAT), grain yield, nutrient content, recovery efficiency, and the amount of N derived from C. ensiformis were quantified. At 54 DAT, the N content, chlorophyll content, and plant height of inoculated plants had increased significantly compared with the control, and these plants were more proficient in the use of N derived from C. ensiformis. At maturation, plants inoculated with A103 were distinguished by the recovery efficiency and amount of N derived from C. ensiformis and N content in the grain and shoot being equal to that in A101 inoculation and higher than that in the control, resulting in a higher accumulation of crude protein and dry matter in the full grain and panicle of DSE-rice interaction. In addition, Fe and Ni contents in the grains of rice inoculated with these fungi doubled with respect to the control, and in A103 inoculation, we observed Mn accumulation that was three times higher than in the other treatments. Our results suggest that the inoculation of rice with DSE fungi represents a strategy to improve green manure-N recovery, grain yield per plant, and grain quality in terms of micronutrients contents in cropping systems with a low N input.

Capsicum annuum L. trypsin inhibitor as a template scaffold for new drug development against pathogenic yeast.

A 6,000 Da peptide, named CaTI, was isolated from Capsicum annuum L. seeds and showed potent inhibitory activity against trypsin and chymotrypsin. The aim of this study was to determine the effect of CaTI on Saccharomyces cerevisiae, Candida albicans, Candida tropicalis and Kluyveromyces marxiannus cells. We observed that CaTI inhibited the growth of S. cerevisiae, K. marxiannus as well as C. albicans and induced cellular agglomeration and the release of cytoplasmic content. No effect on growth was observed in C. tropicalis but morphological changes were noted. In the spot assay, different degrees of sensitivity were shown among the strains and concentrations tested. Scanning electron microscopy showed that S. cerevisiae, K. marxiannus and C. albicans, in the presence of CaTI, exhibited morphological alterations, such as the formation of pseudohyphae, cellular aggregates and elongated forms. We also show that CaTI induces the generation of nitric oxide and interferes in a dose-dependent manner with glucose-stimulated acidification of the medium mediated by H(+)-ATPase of S. cerevisiae cells.

Humic acid effect on catalase activity and the generation of reactive oxygen species in corn (Zea mays).

Humic acids (HAs) have positive effects on plant physiology, but the molecular mechanisms underlying these events are only partially understood. The induction of root growth and emission of lateral roots (LRs) promoted by exogenous auxin is a natural phenomenon. Exogenous auxins are also associated with HA. Gas nitric oxide (NO) is a secondary messenger produced endogenously in plants. It is associated with metabolic events dependent on auxin. With the application of auxin, NO production is significantly increased, resulting in positive effects on plant physiology. Thus it is possible to evaluate the beneficial effects of the application of HA as an effect of auxin. To investigate the effects of HA the parameters of root growth, Zea mays was studied by evaluating the application of 3 mM C L⁻¹ of HA extracted from Oxisol and 100 µM SNP (sodium nitroprusside) and the NO donor, subject to two N-NO3⁻, high dose (5.0 mM N-NO3⁻) and low dose (5.0 mM N-NO3⁻). Treatments with HA and NO were positively increased, regardless of the N-NO3⁻ taken, as assessed by fresh weight and dry root, issue of LRs. The effects were more pronounced in the treatment with a lower dose of N-NO3⁻. Detection of reactive oxygen species (ROS) in vivo and catalase activity were evaluated; these tests were associated with root growth. Under application of the bioactive substances tested, detection of ROS and catalase activity increased, especially in treatments with lower doses of N-NO3⁻. The results of this experiment indicate that the effects of HA are dependent on ROS generation, which act as a messenger that induces root growth and the emission of LRs.