The auxin-responsive CsSPL9-CsGH3.4 module finely regulates auxin levels to suppress the development of adventitious roots in tea (Camellia sinensis).

The cutting technique is extensively used in tea breeding, with key emphasis on promoting the growth of adventitious roots (ARs). Despite its importance in tea cultivation, the mechanisms underlying AR development in tea remain unclear. In this study, we demonstrated the essential role of auxins in the initiation and progression of AR and established that the application of exogenous 1-naphthaleneacetic acid-enhanced AR formation in tissue-cultured seedlings and cuttings. Then, we found that the auxin-responsive transcription factor CsSPL9 acted as a negative regulator of AR development by reducing the levels of free indole-3-acetic acid (IAA) in tea plants. Furthermore, we identified CsGH3.4 as a downstream target of CsSPL9, which was activated by direct binding to its promoter. CsGH3.4 also inhibited AR development and maintained low levels of free IAA. Thus, these results revealed the inhibitory effect of the auxin-responsive CsSPL9-CsGH3.4 module on AR development by reducing free IAA levels in tea. These findings have significant theoretical and practical value for enhancing tea breeding practices.

Exogenous Indole-3-Acetic Acid Suppresses Rice Infection of Magnaporthe oryzae by Affecting Plant Resistance and Fungal Growth.

Auxin is an important phytohormone that regulates diverse biologic processes, including plant growth and immunity. Indole-3-acetic acid (IAA), known as one of the main forms of auxin, is able to activate plant immunity. However, it is unknown whether IAA enhances plant resistance and/or suppresses the growth of the fungal pathogen Magnaporthe oryzae. Here, we found that IAA could induce expression levels of pathogenesis-related genes to enhance disease resistance and could control the development of blast disease through inhibiting M. oryzae infection. Exogenous IAA suppressed mycelial growth and delayed spore germination by inhibiting fungal endogenous IAA biosynthesis and impairing redox homeostasis, respectively. When applied to a field test, two IAA analogues, 1-naphthaleneacetic acid and 2,4-dichlorophenoxy acetic acid, can effectively control rice blast disease. Our study advances the understanding of IAA in controlling rice blast disease through suppressing pathogen growth and enhancing plant resistance.

Improvement of Bioactive Polyphenol Accumulation in Callus of Salvia atropatana Bunge.

Callus cultures of the Iranian medicinal plant Salvia atropatana were initiated from three-week-old seedlings on Murashige and Skoog (MS) medium supplemented with α-naphthaleneacetic acid (NAA) and various cytokinins. Although all tested hormonal variants of the medium and explant enabled callus induction, the most promising growth was noted for N-(2-chloro-4-pyridyl)-N'-phenylurea (CPPU)-induced calli. Three lines obtained on this medium (cotyledon line-CL, hypocotyl line-HL, and root line-RL) were preselected for further studies. Phenolic compounds in the callus tissues were identified using UPLC-MS (ultra-performance liquid chromatography-mass spectrometry) and quantified with HPLC (high-performance liquid chromatography). All lines exhibited intensive growth and contained twelve phenolic acid derivatives, with rosmarinic acid predominating. The cotyledon-derived callus line displayed the highest growth index values and polyphenol content; this was exposed to different light-emitting diodes (LED) for improving biomass accumulation and secondary metabolite yield. Under LED treatments, all callus lines exhibited enhanced RA and total phenolic content compared to fluorescent light, with the highest levels observed for white (48.5-50.2 mg/g dry weight) and blue (51.4-53.9 mg/g dry weight) LEDs. The selected callus demonstrated strong antioxidant potential in vitro based on the 2,2'-azino-bis(3-ethylbenzthiazoline-6-sulfonic acid) (ABTS), 2,2-diphenyl-1-picrylhydrazyl (DPPH), and ferric reducing antioxidant power (FRAP) tests. Our findings confirm that the S. atropatana callus system is suitable for enhanced rosmarinic acid production; the selected optimized culture provide high-quality plant-derived products.

Early defoliation induces auxin redistribution, promoting paradormancy release in pear buds.

Paradormancy of fruit trees occurs in summer and autumn when signals from adjacent organs stimulate buds to develop slowly. This stage has received less attention that the other stages of dormancy, and the underlying mechanism remains uncharacterized. Early defoliation in late summer and early autumn is usually followed by out-of-season blooming in pear (Pyrus spp.), which substantially decreases the number of buds the following spring and negatively affects fruit production. This early bud flush is an example of paradormancy release. Here, we determined that flower bud auxin content is stable after defoliation; however, polar distribution of the pear (Pyrus pyrifolia) PIN-FORMED auxin efflux carrier 1b (PpyPIN1b) implied that auxin tends to be exported from buds. Transcriptome analysis of floral buds after artificial defoliation revealed changes in auxin metabolism, transport, and signal transduction pathways. Exogenous application of a high concentration of the auxin analog 1-naphthaleneacetic acid (300 mg/L) suppressed PpyPIN1b expression and its protein accumulation in the cell membrane, likely leading to decreased auxin efflux from buds, which hindered flower bud sprouting. Furthermore, carbohydrates and additional hormones also influenced out-of-season flowering. Our results indicate that defoliation-induced auxin efflux from buds accelerates bud paradormancy release. This differs from release of apical-dominance-related lateral bud paradormancy after the apex is removed. Our findings and proposed model further elucidate the mechanism underlying paradormancy and will help researchers to develop methods for inhibiting early defoliation-induced out-of-season bud sprouting.

Systematic analysis of specific and nonspecific auxin effects on endocytosis and trafficking.

The phytohormone auxin and its directional transport through tissues are intensively studied. However, a mechanistic understanding of auxin-mediated feedback on endocytosis and polar distribution of PIN auxin transporters remains limited due to contradictory observations and interpretations. Here, we used state-of-the-art methods to reexamine the auxin effects on PIN endocytic trafficking. We used high auxin concentrations or longer treatments versus lower concentrations and shorter treatments of natural indole-3-acetic acid (IAA) and synthetic naphthalene acetic acid (NAA) auxins to distinguish between specific and nonspecific effects. Longer treatments of both auxins interfere with Brefeldin A-mediated intracellular PIN2 accumulation and also with general aggregation of endomembrane compartments. NAA treatment decreased the internalization of the endocytic tracer dye, FM4-64; however, NAA treatment also affected the number, distribution, and compartment identity of the early endosome/trans-Golgi network, rendering the FM4-64 endocytic assays at high NAA concentrations unreliable. To circumvent these nonspecific effects of NAA and IAA affecting the endomembrane system, we opted for alternative approaches visualizing the endocytic events directly at the plasma membrane (PM). Using total internal reflection fluorescence microscopy, we saw no significant effects of IAA or NAA treatments on the incidence and dynamics of clathrin foci, implying that these treatments do not affect the overall endocytosis rate. However, both NAA and IAA at low concentrations rapidly and specifically promoted endocytosis of photo-converted PIN2 from the PM. These analyses identify a specific effect of NAA and IAA on PIN2 endocytosis, thus, contributing to its polarity maintenance and furthermore illustrate that high auxin levels have nonspecific effects on trafficking and endomembrane compartments.

ZmEHD1 Is Required for Kernel Development and Vegetative Growth through Regulating Auxin Homeostasis.

The roles of C-terminal Eps15 homology domain (EHD) proteins in clathrin-mediated endocytosis in plants are poorly understood. Here, we isolated a maize (Zea mays) mutant, designated ehd1, which showed defects in kernel development and vegetative growth. Positional cloning and transgenic analysis revealed that ehd1 encodes an EHD protein. Internalization of the endocytic tracer FM4-64 was substantially reduced in the ehd1 mutant and ZmEHD1 knockout mutants. We further demonstrated that ZmEHD1 and the ZmAP2 σ subunit physically interact at the plasma membrane. Auxin distribution and ZmPIN1a-YFP localization were altered in the ehd1 mutant. Kernel indole-3-acetic acid levels were substantially lower in the ehd1 mutant than in wild-type maize. Exogenous application of 1-naphthaleneacetic acid, but not GA3 or 2-naphthaleneacetic acid, rescued the seed germination and seedling emergency phenotypic defects of ehd1 mutants. Taken together, these results indicate that ZmEHD1 regulates auxin homeostasis by mediating clathrin-mediated endocytosis through its interaction with the ZmAP2 σ subunit, which is crucial for kernel development and vegetative growth of maize.

Role of human flavin-containing monooxygenase (FMO) 5 in the metabolism of nabumetone: Baeyer-Villiger oxidation in the activation of the intermediate metabolite, 3-hydroxy nabumetone, to the active metabolite, 6-methoxy-2-naphthylacetic acid in vitro.

Nabumetone (NAB) is a non-steroidal anti-inflammatory drug used clinically, and its biotransformation includes the major active metabolite 6-methoxy-2-naphthylacetic acid (6-MNA). One of the key intermediates between NAB and 6-MNA may be 3-hydroxy nabumetone (3-OH-NAB). The aim of the present study was to investigate the role of flavin-containing monooxygenase (FMO) isoform 5 in the formation of 6-MNA from 3-OH-NAB. To elucidate the biotransformation of 3-OH-NAB to 6-MNA, an authentic standard of 3-OH-NAB was synthesised and used as a substrate in an incubation with human liver samples or recombinant enzymes. The formation of 3-OH-NAB was observed after the incubation of NAB with various cytochrome P450 (CYP) isoforms. However, 6-MNA itself was rarely detected from NAB and 3-OH-NAB. Further experiments revealed a 6-MNA peak derived from 3-OH-NAB in human hepatocytes. 6-MNA was also detected in the extract obtained from 3-OH-NAB by a combined incubation of recombinant human FMO5 and human liver S9. We herein demonstrated that the reaction involves carbon-carbon cleavage catalyzed by the Baeyer-Villiger oxidation (BVO) of a carbonyl compound, the BVO substrate, such as a ketol, by FMO5. Further in vitro inhibition experiments showed that multiple non-CYP enzymes are involved in the formation of 6-MNA from 3-OH-NAB.

Combined Metabolite and Transcriptome Profiling Reveals the Norisoprenoid Responses in Grape Berries to Abscisic Acid and Synthetic Auxin.

The abscisic acid (ABA) increase and auxin decline are both indicators of ripening initiation in grape berry, and norisoprenoid accumulation also starts at around the onset of ripening. However, the relationship between ABA, auxin, and norisoprenoids remains largely unknown, especially at the transcriptome level. To investigate the transcriptional and posttranscriptional regulation of the ABA and synthetic auxin 1-naphthaleneacetic acid (NAA) on norisoprenoid production, we performed time-series GC-MS and RNA-seq analyses on Vitis vinifera L. cv. Cabernet Sauvignon grape berries from pre-veraison to ripening. Higher levels of free norisoprenoids were found in ABA-treated mature berries in two consecutive seasons, and both free and total norisoprenoids were significantly increased by NAA in one season. The expression pattern of known norisoprenoid-associated genes in all samples and the up-regulation of specific alternative splicing isoforms of VviDXS and VviCRTISO in NAA-treated berries were predicted to contribute to the norisoprenoid accumulation in ABA and NAA-treated berries. Combined weighted gene co-expression network analysis (WGCNA) and DNA affinity purification sequencing (DAP-seq) analysis suggested that VviGATA26, and the previously identified switch genes of myb RADIALIS (VIT_207s0005g02730) and MAD-box (VIT_213s0158g00100) could be potential regulators of norisoprenoid accumulation. The positive effects of ABA on free norisoprenoids and NAA on total norisoprenoid accumulation were revealed in the commercially ripening berries. Since the endogenous ABA and auxin are sensitive to environmental factors, this finding provides new insights to develop viticultural practices for managing norisoprenoids in vineyards in response to changing climates.

Scarlet Flax Linum grandiflorum (L.) In Vitro Cultures as a New Source of Antioxidant and Anti-Inflammatory Lignans.

In vitro cultures of scarlet flax (Linum grandiflorum L.), an important ornamental flax, have been established as a new possible valuable resource of lignans and neolignans for antioxidant and anti-inflammatory applications. The callogenic potential at different concentrations of α-naphthalene acetic acid (NAA) and thidiazuron (TDZ), alone or in combinations, was evaluated using both L. grandiflorum hypocotyl and cotyledon explants. A higher callus induction frequency was observed on NAA than TDZ, especially for hypocotyl explants, with a maximum frequency (i.e., 95.2%) on 1.0 mg/L of NAA. The presence of NAA (1.0 mg/L) in conjunction with TDZ tended to increase the frequency of callogenesis relative to TDZ alone, but never reached the values observed with NAA alone, thereby indicating the lack of synergy between these two plant growth regulators (PGRs). Similarly, in terms of biomass, NAA was more effective than TDZ, with a maximum accumulation of biomass registered for medium supplemented with 1.0 mg/L of NAA using hypocotyls as initial explants (DW: 13.1 g). However, for biomass, a synergy between the two PGRs was observed, particularly for cotyledon-derived explants and for the lowest concentrations of TDZ. The influence of these two PGRs on callogenesis and biomass is discussed. The HPLC analysis confirmed the presence of lignans (secoisolariciresinol (SECO) and lariciresinol (LARI) and neolignan (dehydrodiconiferyl alcohol [DCA]) naturally accumulated in their glycoside forms. Furthermore, the antioxidant activities performed for both hypocotyl- and cotyledon-derived cultures were also found maximal (DPPH: 89.5%, FRAP 866: µM TEAC, ABTS: 456 µM TEAC) in hypocotyl-derived callus cultures as compared with callus obtained from cotyledon explants. Moreover, the anti-inflammatory activities revealed high inhibition (COX-1: 47.4% and COX-2: 51.1%) for extract of hypocotyl-derived callus cultures at 2.5 mg/L TDZ. The anti-inflammatory action against COX-1 and COX-2 was supported by the IC50 values. This report provides a viable approach for enhanced biomass accumulation and efficient production of (neo)lignans in L. grandiflorum callus cultures.

Auxin treatment of grapevine (Vitis vinifera L.) berries delays ripening onset by inhibiting cell expansion.

KEY MESSAGE: Auxin treatment of grape (Vitis vinifera L.) berries delays ripening by inducing changes in gene expression and cell wall metabolism and could combat some deleterious climate change effects. Auxins are inhibitors of grape berry ripening and their application may be useful to delay harvest to counter effects of climate change. However, little is known about how this delay occurs. The expression of 1892 genes was significantly changed compared to the control during a 48 h time-course where the auxin 1-naphthaleneacetic acid (NAA) was applied to pre-veraison grape berries. Principal component analysis showed that the control and auxin-treated samples were most different at 3 h post-treatment when approximately three times more genes were induced than repressed by NAA. There was considerable cross-talk between hormone pathways, particularly between those of auxin and ethylene. Decreased expression of genes encoding putative cell wall catabolic enzymes (including those involved with pectin) and increased expression of putative cellulose synthases indicated that auxins may preserve cell wall structure. This was confirmed by immunochemical labelling of berry sections using antibodies that detect homogalacturonan (LM19) and methyl-esterified homogalacturonan (LM20) and by labelling with the CMB3a cellulose-binding module. Comparison of the auxin-induced changes in gene expression with the pattern of these genes during berry ripening showed that the effect on transcription is a mix of changes that may specifically alter the progress of berry development in a targeted manner and others that could be considered as non-specific changes. Several lines of evidence suggest that cell wall changes and associated berry softening are the first steps in ripening and that delaying cell expansion can delay ripening providing a possible mechanism for the observed auxin effects.

Optimization of growth regulators on in vitro propagation of Moringa stenopetala from shoot explants.

BACKGROUND: Moringa stenopetala belongs to the flowering family Moringaceae and genus Moringa. It is often referred to as the East African Moringa tree because it is native only to southern Ethiopia and northern Kenya. The expansion of its cultivation and utilization throughout the world especially in Africa is becoming important. For such expansion, the existing propagation method is limiting, so it needs a good propagation system to supply enough planting material with a uniform genotype. Therefore, the main objective of this study was to optimize an in vitro shoot multiplication protocol for M. stenopetala by using shoot tip as explants. RESULTS: Shoots were sterilized and cultured on Muraghige and Skoog (MS) medium for in vitro shoot initiation. For multiple shoot induction, the explants were cultured on MS medium supplemented with different concentrations of kinetin (0.5, 1.0, 1.5, 2.0, 2.5 mg/L) with Indole-3- butyric acid (IBA) or α -naphthalene acetic acid (NAA) (0.01, 0.1, 0.5 mg/L) and maintained at 25 ± 2 °C for four weeks. Rooting was achieved by culturing well developed shoots in half-strength MS medium containing IBA (0.1, 0.5, 1.0, 1.5, 2.0 mg/L), NAA (0.1, 0.5, 1.0, 1.5, 2.0 mg/L), and 0.5 mg/L IBA with NAA (0.1, 0.5, 1.0, 1.5, 2.0 mg/L). Statistical analysis revealed that there was a significant difference among all treatments applied in both shoot multiplication and rooting experiments. The maximum number of shoots per explant (3.43 ± 1.41) and 7.97 ± 4.18 leaves per explant were obtained on MS medium containing 0.5 mg/L kinetin with 0.01 mg/LNAA. The highest mean number of roots per shoot (1.63 ± 1.03) and mean root length (0.87 ± 1.22 cm) were obtained on MS medium containing 1.0 mg/LNAA and 0.1 mg/LIBA alone respectively. After acclimatization, 76% of plants were survived in the greenhouse. CONCLUSION: In general, using NAA with kinetin for shoot multiplication was effective than kinetin with IBA. On the other hand, the application of 1.0 mg/L NAA alone and 1.0 mg/L NAA with 0.5 mg/L IBA were more effective for root induction.

Indirect somatic embryogenesis of purple coneflower (Echinacea purpurea (L.) Moench): a medicinal-ornamental plant: evaluation of antioxidant enzymes activity and histological study.

Purple coneflower (Echinacea purpurea (L.) Moench) is a widely used medicinal and ornamental plant. In the present study, the callus embryogenesis was examined using benzyl adenine (BA) at three levels (3, 4, 5 mg L-1), 1-Naphthalene acetic acid (NAA) at three levels (0.1, 0.2 and 0.5 mg L-1) with or without activated charcoal (1 g L-1), coconut milk (50 ml L-1) and casein hydrolysate (50 mg L-1) in the MS (Murashige and Skoog 1962) medium. The embryogenesis indirectly occurred with the production of callus. The calli were observed in three forms: undifferentiated, embryogenic and organogenic. The embryogenic calli were dark green and coherent with a faster growth rate. The highest embryogenesis (100%) and embryonic regeneration (plantlet production) were obtained in the combined BA + NAA treatments with the activated charcoal, coconut milk and casein hydrolysate. However, the combined treatments of growth regulators failed to produce somatic embryos without the use of coconut milk and casein hydrolysate. The maximum amount of protein, peroxidase and catalase activity of embryogenic calli (2.02, 1.79 and 6.62ΔOD/Min/mg.protein, respectively), and highest percentage of acclimatization success (29.3% of plants) were obtained in the combined treatment of 5 mg L-1 BA + 0.5 mg L-1 NAA + activated charcoal + coconut milk + casein hydrolysate. The highest amount of chlorophyll content (33.3 SPAD value) and growth characteristics of acclimatized plantlets were observed in the media containing 3 mg L-1 BA + 0.1 and 0.2 mg L-1 NAA + 1 g. L-1 combined activated charcoal, coconut milk, casein hydrolysate. The histological studies confirmed the somatic embryogenesis in purple coneflower. Generally, it was found that the somatic embryogenesis of E. purpurea occurs at high levels of BA and low levels of NAA with the addition of coconut milk and casein hydrolysate.

Establishment of an Agrobacterium mediated transformation protocol for the detection of cytokinin in the heterophyllous plant Hygrophila difformis (Acanthaceae).

KEY MESSAGE: This is the first report of a highly efficient Agrobacterium tumefaciens-mediated transformation protocol for Acanthaceae and its utilization in revealing important roles of cytokinin in regulating heterophylly in Hygrophila difformis. Plants show amazing morphological differences in leaf form in response to changes in the surrounding environment, which is a phenomenon called heterophylly. Previous studies have shown that the aquatic plant Hygrophila difformis (Acanthaceae) is an ideal model for heterophylly study. However, low efficiency and poor reproducibility of genetic transformation restricted H. difformis as a model plant. In this study, we reported successful induction of callus, shoots and the establishment of an efficient stable transformation protocol as mediated by Agrobacterium tumefaciens LBA4404. We found that the highest callus induction efficiency was achieved with 1 mg/L 1-Naphthaleneacetic acid (NAA) and 2 mg/L 6-benzyladenine (6-BA), that efficient shoot induction required 0.1 mg/L NAA and 0.1 mg/L 6-BA and that high transformation efficiency required 100 µM acetosyringone. Due to the importance of phytohormones in the regulation of heterophylly and the inadequate knowledge about the function of cytokinin (CK) in this process, we analyzed the function of CK in the regulation of heterophylly by exogenous CK application and endogenous CK detection. By using our newly developed transformation system to detect CK signals, contents and distribution in H. difformis, we revealed an important role of CK in environmental mediated heterophylly.

A metabolic pathway for the prodrug nabumetone to the pharmacologically active metabolite, 6-methoxy-2-naphthylacetic acid (6-MNA) by non-cytochrome P450 enzymes.

The pathway for the transformation of the prodrug nabumetone, 4-(6-methoxynaphthalen-2-yl)butan-2-one, to the active metabolite 6-methoxy-2-naphthylacetic acid (6-MNA), a potent cyclooxygenase-2 inhibitor, has not yet been clarified in humans.To confirm the activation pathway, authentic standards of the nabumetone intermediates, 2-(6-methoxynaphthalen-2-yl)ethyl acetate (6-MNEA), 2-(6-methoxynaphthalen-2-yl)ethan-1-ol (6-MNE-ol) and 2-(6-methoxynaphthalen-2-yl)acetaldehyde (6-MN-CHO) were synthesized. High performance liquid-chromatography and gas chromatography-mass spectrometry on nabumetone oxidation revealed the generation of three metabolites.The formation of 6-MNA after a 60-min incubation of nabumetone was detected and 6-MNE-ol, an alcohol-related intermediate, was also generated by in cryopreserved hepatocytes. However, 6-MNA was below detection limit, but 4-(6-methoxynaphthalen-2-yl)butan-2-ol (MNBO) and 4-(6-hydroxynaphthalen-2-yl)butan-2-one (M3) peak were found in both the microsomes and S9 extracts with any cofactors.Nabumetone has recently been proposed as a typical substrate of flavin-containing monooxygenase isoform 5 (FMO5) and was shown to be efficiently oxidized in vitro to 6-MNEA. 6-MNA was detected in the extract obtained from a combined incubation of recombinant FMO5 and S9 fractions.The specificity of FMO5 towards catalyzing this Baeyer-Villiger oxidation (BVO) was demonstrated by the inhibition of the BVO substrate, 4-methoxyphenylacetone. Further in vitro inhibition studies demonstrated that multiple non-cytochrome P450 enzymes are involved in the formation of 6-MNA.

In Vitro Micropropagation of Industrially and Medicinally Useful Plant Aloe trichosantha Berger Using Offshoot Cuttings.

This study was aimed to develop in vitro micropropagation protocol of Aloe trichosantha Berger using offshoots as explants. MS media supplemented with plant growth regulators helped explants develop shoots within about 14 to 17 days. The mean number of days to shooting has decreased from 16.8 ± 0.8 with 0.5/0.5 mg/L BAP/NAA supplement to 15.5 ± 0.5 with 2.0/0.5 mg/L BAP/NAA. While the mean shoot number has increased with increasing the concentration of BAP supplements, the reverse was true with mean shoot lengths, whereas supplement of 2.0/0.5 mg/L BAP/NAA has generated significantly more shoots (17 ± 3.8), and longer shoots were produced with the addition of 0.5/0.5 and 1.0/0.5 mg/L BAP/NAA. In regard to rooting, though higher concentrations of NAA have resulted in quick rooting, the rooting performance in terms of mean number and length of roots was better with low concentrations. All the plantlets subjected to greenhouse acclimatization in cocopeat have survived. Secondary acclimatization in composted and manured soil media has also resulted in 93 to 95% survival rate. Lighting conditions (nursery shade or direct sunlight) of secondary acclimatization did not lead to any difference in the survival rate of the plantlets.

Different Roles of Auxins in Somatic Embryogenesis Efficiency in Two Picea Species.

The effects of auxins 2,4-D (2,4-dichlorophenoxyacetic acid), NAA (1-naphthaleneacetic acid) or picloram (4-amino-3,5,6-trichloropicolinic acid; 9 µM) and cytokinin BA (benzyloadenine; 4.5 µM) applied in the early stages of somatic embryogenesis (SE) on specific stages of SE in Picea abies and P. omorika were investigated. The highest SE initiation frequency was obtained after 2,4-D application in P. omorika (22.00%) and picloram application in P. abies (10.48%). NAA treatment significantly promoted embryogenic tissue (ET) proliferation in P. abies, while 2,4-D treatment reduced it. This reduction was related to the oxidative stress level, which was lower with the presence of NAA in the proliferation medium and higher with the presence of 2,4-D. The reduced oxidative stress level after NAA treatment suggests that hydrogen peroxide (H2O2) acts as a signalling molecule and promotes ET proliferation. NAA and picloram in the proliferation medium decreased the further production and maturation of P. omorika somatic embryos compared with that under 2,4-D. The quality of the germinated P. abies embryos and their development into plantlets depended on the auxin type and were the highest in NAA-originated embryos. These results show that different auxin types can generate different physiological responses in plant materials during SE in both spruce species.

Colonization of endophyte Acremonium sp. D212 in Panax notoginseng and rice mediated by auxin and jasmonic acid.

Endophytic fungi can be beneficial to plant growth. However, the molecular mechanisms underlying colonization of Acremonium spp. remain unclear. In this study, a novel endophytic Acremonium strain was isolated from the buds of Panax notoginseng and named Acremonium sp. D212. The Acremonium sp. D212 could colonize the roots of P. notoginseng, enhance the resistance of P. notoginseng to root rot disease, and promote root growth and saponin biosynthesis in P. notoginseng. Acremonium sp. D212 could secrete indole-3-acetic acid (IAA) and jasmonic acid (JA), and inoculation with the fungus increased the endogenous levels of IAA and JA in P. notoginseng. Colonization of the Acremonium sp. D212 in the roots of the rice line Nipponbare was dependent on the concentration of methyl jasmonate (MeJA) (2-15 µmol/L) and 1-naphthalenacetic acid (NAA) (10-20 µmol/L). Moreover, the roots of the JA signaling-defective coi1-18 mutant were colonized by Acremonium sp. D212 to a lesser degree than those of the wild-type Nipponbare and miR393b-overexpressing lines, and the colonization was rescued by MeJA but not by NAA. It suggests that the cross-talk between JA signaling and the auxin biosynthetic pathway plays a crucial role in the colonization of Acremonium sp. D212 in host plants.

Auxin Efflux Carrier ZmPGP1 Mediates Root Growth Inhibition under Aluminum Stress.

Auxin has been shown to enhance root growth inhibition under aluminum (Al) stress in Arabidopsis (Arabidopsis thaliana). However, in maize (Zea mays), auxin may play a negative role in the Al-induced inhibition of root growth. In this study, we identified mutants deficient in the maize auxin efflux carrier P-glycoprotein (ZmPGP1) after ethyl methanesulfonate mutagenesis and used them to elucidate the contribution of ZmPGP1 to Al-induced root growth inhibition. Root growth in the zmpgp1 mutant, which forms shortened roots and is hyposensitive to auxin, was less inhibited by Al stress than that in the inbred line B73. In the zmpgp1 mutants, the root tips displayed higher auxin accumulation and enhanced auxin signaling under Al stress, which was also consistent with the increased expression of auxin-responsive genes. Based on the behavior of the auxin-responsive marker transgene, DR5rev:RFP, we concluded that Al stress reduced the level of auxin in the root tip, which contrasts with the tendency of Al stress-induced Arabidopsis plants to accumulate more auxin in their root tips. In addition, Al stress induced the expression of ZmPGP1 Therefore, in maize, Al stress is associated with reduced auxin accumulation in root tips, a process that is regulated by ZmPGP1 and thus causes inhibition of root growth. This study provides further evidence about the role of auxin and auxin polar transport in Al-induced root growth regulation in maize.

Phenolic compounds, antioxidant capacity and antimutagenic activity in different growth stages of in vitro raised plants of Origanum vulgare L.

Efficient micropropagation procedure was developed for Origanum vulgare, a high-value culinary herb, and the phytochemicals, phenolic content, antioxidant and antimutagenic activity of leaf and stem, derived from different growing stages were analyzed. The agar solidified Murashige and Skoog (MS) medium supplemented with a combination of 6-benzylaminopurine and α-naphthaleneacetic acid was optimized as best shoot-multiplication-medium. Shoots were rooted best on 1/2 strength MS medium supplemented with 50 µM indole-3-butyric acid (IBA). The plantlets were successfully acclimatized ex vitro in a soil, sand and farmyard manure mixture (2:1:1 v/v/v) with 100% survival rate in greenhouse. The total anthocyanin and total phenolic content were observed significantly higher in leaves of in vitro-raised plants. However, total tannin, flavonoid and antioxidant activity remained higher in leaves of mother plant maintained under ployhouse condition. All the plant extracts have shown significant antimutagenic activity except in vitro-growing plants. A total of 13 polyphenolic compounds were detected in different extracts using high performance liquid chromatography. Among these, catechin was detected maximum in in vitro-growing cultures and chlorogenic acid in leaves of mother plant. These findings will help the farmers, medicinal plant growers, and industries for mass multiplication and effective extraction of phytochemicals from O. vulgare.

Gut microbiota metabolizes nabumetone in vitro: Consequences for its bioavailability in vivo in the rodents with altered gut microbiome.

1. The underlying microbial metabolic activity toward xenobiotics is among the least explored factors contributing to the inter-individual variability in drug response. 2. Here, we analyzed the effect of microbiota on a non-steroidal anti-inflammatory drug nabumetone. 3. First, we cultivated the drug with the selected gut commensal and probiotic bacteria under both aerobic and anaerobic conditions and analyzed its metabolites by high-performance liquid chromatography (HPLC) with UV detection. To analyze the effect of microbiota on nabumetone pharmacokinetics in vivo, we administered a single oral dose of nabumetone to rodents with intentionally altered gut microbiome - either rats treated for three days with the antibiotic imipenem or to germ-free mice. Plasma levels of its main active metabolite 6 methoxy-2-naphthylacetic acid (6-MNA) were analyzed at pre-specified time intervals using HPLC with UV/fluorescence detection. 4. We found that nabumetone is metabolized by bacteria to its non-active metabolites and that this effect is stronger under anaerobic conditions. Although in vivo, none of the pharmacokinetic parameters of 6-MNA was significantly altered, there was a clear trend towards an increase of the AUC, Cmax and t1/2 in rats with reduced microbiota and germ-free mice.