Studies of moss reproductive development indicate that auxin biosynthesis in apical stem cells may constitute an ancestral function for focal growth control

The plant hormone auxin is a key factor for regulation of plant development, and this function was probably reinforced during the evolution of early land plants. We have extended the available toolbox to allow detailed studies of how auxin biosynthesis and responses are regulated in moss reproductive organs, their stem cells and gametes to better elucidate the function of auxin in the morphogenesis of early land plants. We measured auxin metabolites and identified IPyA (indole-3-pyruvic acid) as the main biosynthesis pathway inPhyscomitrium(Physcomitrella)patensand established knock-out, overexpressor and reporter lines for biosynthesis genes which were analyzed alongside previously reported auxin-sensing and transport reporters. Vegetative and reproductive apical stem cells synthesize auxin. Sustained stem cell activity depends on an inability to sense the auxin produced while progeny of the stem cells respond to the auxin, aiding in the control of cell division, expansion and differentiation. Gamete precursors are dependent on a certain degree of auxin sensing, while the final differentiation is a low auxin-sensing process. Tha data presented indicate that low auxin activity may represent a conserved hallmark of land plant gametes, and that local auxin biosynthesis in apical stem cells may be part of an ancestral mechanism to control focal growth

Inactivation of the entire Arabidopsis group II GH3s confers tolerance to salinity and water deficit

Indole-3-acetic acid (IAA) controls a plethora of developmental processes. Thus, regulation of its concentration is of great relevance for plant performance. Cellular IAA concentration depends on its transport, biosynthesis and the various pathways for IAA inactivation, including oxidation and conjugation. Group II members of the GRETCHEN HAGEN 3 (GH3) gene family code for acyl acid amido synthetases catalysing the conjugation of IAA to amino acids. However, the high degree of functional redundancy among them has hampered thorough analysis of their roles in plant development. In this work, we generated an Arabidopsis gh3.1,2,3,4,5,6,9,17 (gh3oct) mutant to knock out the group II GH3 pathway. The gh3oct plants had an elaborated root architecture, showed an increased tolerance to different osmotic stresses, including an IAA-dependent tolerance to salinity, and were more tolerant to water deficit. Indole-3-acetic acid metabolite quantification in gh3oct plants suggested the existence of additional GH3-like enzymes in IAA metabolism. Moreover, our data suggested that 2-oxindole-3-acetic acid production depends, at least in part, on the GH3 pathway. Targeted stress-hormone analysis further suggested involvement of abscisic acid in the differential response to salinity of gh3oct plants. Taken together, our data provide new insights into the roles of group II GH3s in IAA metabolism and hormone-regulated plant development

An ectomycorrhizal fungus alters sensitivity to jasmonate, salicylate, gibberellin, and ethylene in host roots.

The phytohormones jasmonate, gibberellin, salicylate, and ethylene regulate an interconnected reprogramming network integrating root development with plant responses against microbes. The establishment of mutualistic ectomycorrhizal symbiosis requires the suppression of plant defense responses against fungi as well as the modification of root architecture and cortical cell wall properties. Here, we investigated the contribution of phytohormones and their crosstalk to the ontogenesis of ectomycorrhizae (ECM) between grey poplar (Populus tremula x alba) roots and the fungus Laccaria bicolor. To obtain the hormonal blueprint of developing ECM, we quantified the concentrations of jasmonates, gibberellins, and salicylate via liquid chromatography-tandem mass spectrometry. Subsequently, we assessed root architecture, mycorrhizal morphology, and gene expression levels (RNA sequencing) in phytohormone-treated poplar lateral roots in the presence or absence of L. bicolor. Salicylic acid accumulated in mid-stage ECM. Exogenous phytohormone treatment affected the fungal colonization rate and/or frequency of Hartig net formation. Colonized lateral roots displayed diminished responsiveness to jasmonate but regulated some genes, implicated in defense and cell wall remodelling, that were specifically differentially expressed after jasmonate treatment. Responses to salicylate, gibberellin, and ethylene were enhanced in ECM. The dynamics of phytohormone accumulation and response suggest that jasmonate, gibberellin, salicylate, and ethylene signalling play multifaceted roles in poplar L. bicolor ectomycorrhizal development

Determination of pefloxacin by the technique of sequential injection analysis with chemiluminescence detection

Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Consultant: doc. RNDr. Miroslav Polášek, CSc. Diploma Thesis Title: Determination of pefloxacin mesylate dihydrate using sequential injection analysis with chemiluminescent detection A simple and rapid method for determination of pefloxacin mesylate dihydrate (PEF) using sequential injection analysis with chemiluminescent detection was devised and optimised. Chemiluminescent reaction was based on redox reaction of tris(2,2′- bipyridine)dichlororuthenium(II) hexahydrate. Ammonium cerium(IV) sulfate was used as an oxidant. Sodium acetate was used as a third component which had enhancing effect on CL reaction. The calibration curve was linear in the range 5,2* 10-07 - 1,29* 10-05 M of PEF, R2= 0,9948. The limit of detection (LOD) was 0,21 μM and the limit of quantification (LOQ) was 0,28 μM. This optimised method was used for the determination of PEF in ABAKTAL inj. sol. Interferences from some excipients (EDTA, ascorbic acid and sodium metabisulfite) used in the production of injection solutions were examined

Determination of pefloxacin by the technique of sequential injection analysis with chemiluminescence detection

Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Analytical Chemistry Consultant: doc. RNDr. Miroslav Polášek, CSc. Diploma Thesis Title: Determination of pefloxacin mesylate dihydrate using sequential injection analysis with chemiluminescent detection A simple and rapid method for determination of pefloxacin mesylate dihydrate (PEF) using sequential injection analysis with chemiluminescent detection was devised and optimised. Chemiluminescent reaction was based on redox reaction of tris(2,2′- bipyridine)dichlororuthenium(II) hexahydrate. Ammonium cerium(IV) sulfate was used as an oxidant. Sodium acetate was used as a third component which had enhancing effect on CL reaction. The calibration curve was linear in the range 5,2* 10-07 - 1,29* 10-05 M of PEF, R2= 0,9948. The limit of detection (LOD) was 0,21 μM and the limit of quantification (LOQ) was 0,28 μM. This optimised method was used for the determination of PEF in ABAKTAL inj. sol. Interferences from some excipients (EDTA, ascorbic acid and sodium metabisulfite) used in the production of injection solutions were examined

The RPN12a proteasome subunit is essential for the multiple hormonal homeostasis controlling the progression of leaf senescence

The 26S proteasome is a conserved multi-subunit machinery in eukaryotes. It selectively degrades ubiquitinated proteins, which in turn provides an efficient molecular mechanism to regulate numerous cellular functions and developmental processes. Here, we studied a new loss-of-function allele of RPN12a, a plant ortholog of the yeast and human structural component of the 19S proteasome RPN12. Combining a set of biochemical and molecular approaches, we confirmed that a rpn12a knock-out had exacerbated 20S and impaired 26S activities. The altered proteasomal activity led to a pleiotropic phenotype affecting both the vegetative growth and reproductive phase of the plant, including a striking repression of leaf senescence associate cell-death. Further investigation demonstrated that RPN12a is involved in the regulation of several conjugates associated with the auxin, cytokinin, ethylene and jasmonic acid homeostasis. Such enhanced aptitude of plant cells for survival in rpn12a contrasts with reports on animals, where 26S proteasome mutants generally show an accelerated cell death phenotype

GOLVEN peptides regulate lateral root spacing as part of a negative feedback loop on the establishment of auxin maxima

Lateral root initiation requires the accumulation of auxin in lateral root founder cells, yielding a local auxin maximum. The positioning of auxin maxima along the primary root determines the density and spacing of lateral roots. The GOLVEN6 (GLV6) and GLV10 signaling peptides and their receptors have been established as regulators of lateral root spacing via their inhibitory effect on lateral root initiation in Arabidopsis. However, it remained unclear how these GLV peptides interfere with auxin signaling or homeostasis. Here, we show that GLV6/10 signaling regulates the expression of a subset of auxin response genes, downstream of the canonical auxin signaling pathway, while simultaneously inhibiting the establishment of auxin maxima within xylem-pole pericycle cells that neighbor lateral root initiation sites. We present genetic evidence that this inhibitory effect relies on the activity of the PIN3 and PIN7 auxin export proteins. Furthermore, GLV6/10 peptide signaling was found to enhance PIN7 abundance in the plasma membranes of xylem-pole pericycle cells, which likely stimulates auxin efflux from these cells. Based on these findings, we propose a model in which the GLV6/10 signaling pathway serves as a negative feedback mechanism that contributes to the robust patterning of auxin maxima along the primary root

Salicylic acid metabolism and signalling coordinate senescence initiation in aspen in nature

Abstract Deciduous trees exhibit a spectacular phenomenon of autumn senescence driven by the seasonality of their growth environment, yet there is no consensus which external or internal cues trigger it. Senescence starts at different times in European aspen (Populus tremula L.) genotypes grown in same location. By integrating omics studies, we demonstrate that aspen genotypes utilize similar transcriptional cascades and metabolic cues to initiate senescence, but at different times during autumn. The timing of autumn senescence initiation appeared to be controlled by two consecutive “switches”; 1) first the environmental variation induced the rewiring of the transcriptional network, stress signalling pathways and metabolic perturbations and 2) the start of senescence process was defined by the ability of the genotype to activate and sustain stress tolerance mechanisms mediated by salicylic acid. We propose that salicylic acid represses the onset of leaf senescence in stressful natural conditions, rather than promoting it as often observed in annual plants

Changes in cell wall composition due to a pectin biosynthesis enzyme GAUT10 impact root growth

Arabidopsis (Arabidopsis thaliana) root development is regulated by multiple dynamic growth cues that require central metabolism pathways such as β-oxidation and auxin. Loss of the pectin biosynthesizing enzyme GALACTURONOSYLTRANSFERASE 10 (GAUT10) leads to a short-root phenotype under sucrose-limited conditions. The present study focused on determining the specific contributions of GAUT10 to pectin composition in primary roots and the underlying defects associated with gaut10 roots. Using live-cell microscopy, we determined reduced root growth in gaut10 is due to a reduction in both root apical meristem size and epidermal cell elongation. In addition, GAUT10 was required for normal pectin and hemicellulose composition in primary Arabidopsis roots. Specifically, loss of GAUT10 led to a reduction in galacturonic acid and xylose in root cell walls and altered the presence of rhamnogalacturonan-I (RG-I) and homogalacturonan (HG) polymers in the root. Transcriptomic analysis of gaut10 roots compared to wild type uncovered hundreds of genes differentially expressed in the mutant, including genes related to auxin metabolism and peroxisome function. Consistent with these results, both auxin signaling and metabolism were modified in gaut10 roots. The sucrose-dependent short-root phenotype in gaut10 was linked to β-oxidation based on hypersensitivity to indole-3-butyric acid (IBA) and an epistatic interaction with TRANSPORTER OF IBA1 (TOB1). Altogether, these data support a growing body of evidence suggesting that pectin composition may influence auxin pathways and peroxisome activity.This article is published as Linkan Dash, Sivakumar Swaminathan, Jan Šimura, Caitlin Leigh P Gonzales, Christian Montes, Neel Solanki, Ludvin Mejia, Karin Ljung, Olga A Zabotina, Dior R Kelley, Changes in cell wall composition due to a pectin biosynthesis enzyme GAUT10 impact root growth, Plant Physiology, Volume 193, Issue 4, 2023, Pages 2480–2497, https://doi.org/10.1093/plphys/kiad465. © The Author(s) 2023.This is an Open Access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited