Effects of Nutrient Limitation and {gamma}-Irradiation on Tracheary Element Differentiation and Cell Division in Single Mesophyll Cells of Zinnia elegans

The effects of nutrient limitation and -irradiation on trachearyelement differentiation and cell division were investigatedusing single cells isolated from the mesophyll of Zinnia elegans.When the phosphate concentration of the medium was reduced to10 µM (1/50 of Fukuda and Komamine's medium, 1980a), thefrequency of cell division during 4 days of culture decreased,while the frequency of tracheary element differentiation wasunaffected. -Irradiation with a dose of 92 Gy at 36 h of culturepreferentially and thoroughly suppressed cell division withoutreducing the number of tracheary elements formed. The appearanceof secondary cell wall thickenings was delayed by irradiation,but synchrony was maintained. Thus the Zinnia system previouslyreported [Fukuda and Komamine (1980a) Plant Physiol. 65: 57]was improved to give a more useful system for the study of cytodifferentiation,in which tracheary element formation occurred from single cellswithout cell division. ¹Present address: Biological Institute, Faculty of Science,Tohoku University, Sendai, Miyagi 980, Japan. (Received November 28, 1985; Accepted February 22, 1986)     

Interrelationship between lignin deposition and the activities of peroxidase isoenzymes in differentiating tracheary elements of Zinnia

In a culture system in which single cells isolated from the mesophyll of Zinnia elegans L. differentiate to tracheary elements (TEs), two inhibitors of phenylalanine ammonia-lyase (EC 4.3.1.5), L-α-aminooxy-β-phenylpropionic acid (AOPP) at 10 μM inhibited lignification without reducing the number of TEs formed. These inhibitors caused intracellular changes in peroxidase (EC 1.11.1.7) activities. The inhibitors increased the activity of peroxidases bound to the cell walls and especially the activity of peroxidase bound ionically to the cell walls. In contrast, the activity of extracellular peroxidase decreased. There were five isoenzymes, P1-P5, in the ionically bound peroxidase of cultured Zinnia cells. Among the isoenzymes, P4 and P5 appeared to be specific for TE differentation. Treatment with AOPP and AIP resulted in increases in the activities of P2, P4 and P5 isoenzymes, with the most prominent increase in P5 activity. The addition of lignin precursors, including coniferyl alcohol, to the AOPP-treated cells restored lignification, and suppressed the alteration of peroxidase isoenzyme patterns caused by AOPP. The relationship between the wall-bound peroxidases and lignification during TE differentiation is discussed in the light of these results.     

Identification of a multi-component berberine 11-hydroxylase from Burkholderia sp. strain CJ1

ABSTRACT

Berberine (BBR) is a protoberberine alkaloid extracted from plants such as Coptis japonica (Ranunculaceae). In a previous report, we demonstrated the existence of a 11-hydroxylation pathway employed by BBR-utilizing bacteria for metabolism of BBR. In the present study, we report the identification of the genes brhA, brhB, and brhC as encoding a multicomponent BBR 11-hydroxylase in Burkholderia sp. strain CJ1. BrhA is belonging to the Rieske non-heme iron oxygenase (RO) family, a class of enzymes known to catalyze the first step in bacterial aromatic-ring hydroxylation. We further demonstrate that BrhA activity requires BrhB (ferredoxin reductase) and BrhC (ferredoxin) as electron transport chain components. A BLAST search revealed that BrhA exhibits 38% and 33% sequence identity to dicamba O-demethylase (DdmC; AY786443) and chloroacetanilide herbicides N-dealkylase (CndA; KJ461679), respectively. To our knowledge, this work represents the first report of a bacterial oxygenase catalyzing the metabolism of a polycyclic aromatic-ring alkaloid.

Abbreviations: BBR: berberine; D-BBR: demethyleneberberine; H-BBR: 11-hydroxyberberine; HD-BBR: 11-hydroxydemethyleneberberine; HDBA: 2-hydroxy-3,4-dimethoxybenzeneacetic acid; PAL: palmatine; H-PAL: 11-hydroxypalmatine; BRU: berberrubine; Fd: ferredoxin; FdR: ferredoxin reductase; ETC: electron transport chain     

GH–IGF system regulation of attenuated muscle growth and lipolysis in Atlantic salmon reared at elevated sea temperatures

Growth regulation in adult Atlantic salmon (1.6 kg) was investigated during 45 days in seawater at 13, 15, 17, and 19 °C. We focused on feed intake, nutrient uptake, nutrient utilization, and endocrine regulation through growth hormone (GH), insulin-like growth factors (IGF), and IGF-binding proteins (IGFBP). During prolonged thermal exposure, salmon reduced feed intake and growth. Feed utilization was reduced at 19 °C after 45 days compared with fish at lower temperatures, and body lipid storage was depleted with increasing water temperature. Although plasma IGF-1 concentrations did not change, 32-Da and 43-kDa IGFBP increased in fish reared at ≤17 °C, and dropped in fish reared at 19 °C. Muscle igf1 mRNA levels were reduced at 15 and 45 days in fish reared at 15, 17, and 19 °C. Muscle igf2 mRNA levels did not change after 15 days in response to increasing temperature, but were reduced after 45 days. Although liver igf2 mRNA levels were reduced with increasing temperatures after 15 and 45 days, temperature had no effect on igf1 mRNA levels. The liver igfbp2b mRNA level, which corresponds to circulating 43-kDa IGFBP, exhibited similar responses after 45 days. IGFBP of 23 kDa was only detected in plasma in fish reared at 17 °C, and up-regulation of the corresponding igfbp1b gene indicated a time-dependent catabolic response, which was not observed in fish reared at 19 °C. However, higher muscle ghr mRNA levels were detected in fish at 17 and 19 °C than in fish at lower temperatures, indicating lipolytic regulation in muscle. These results show that the reduction of muscle growth in large salmon is mediated by decreased igf1 and igf2 mRNA levels in addition to GH-associated lipolytic action to cope with prolonged thermal exposure. Accordingly, 13 °C appears to be a more optimal temperature for the growth of adult Atlantic salmon at sea.     

Discovery of a small-molecule inhibitor and cellular probe of Keap1–Nrf2 protein–protein interaction

A high-throughput screen (HTS) of the MLPCN library using a homogenous fluorescence polarization assay identified a small molecule as a first-in-class direct inhibitor of Keap1–Nrf2 protein–protein interaction. The HTS hit has three chiral centers; a combination of flash and chiral chromatographic separation demonstrated that Keap1-binding activity resides predominantly in one stereoisomer (SRS)-5 designated as ML334 (LH601A), which is at least 100× more potent than the other stereoisomers. The stereochemistry of the four cis isomers was assigned using X-ray crystallography and confirmed using stereospecific synthesis. (SRS)-5 is functionally active in both an ARE gene reporter assay and an Nrf2 nuclear translocation assay. The stereospecific nature of binding between (SRS)-5 and Keap1 as well as the preliminary but tractable structure–activity relationships support its use as a lead for our ongoing optimization     

Evaluation of bifidobacterial adhesion to acidic sugar chains of porcine colonic mucins

The aim of this study was to assess the adhesion of Bifidobacterium strains to acidic carbohydrate moieties of porcine colonic mucin. Mucins were extracted and purified via gel filtration chromatography followed by density-gradient ultracentrifugation. The presence of sulfated and sialylated carbohydrates in mucins was shown by enzyme-linked immunosorbent assays using PGM34 and HMC31 monoclonal antibodies (mAbs), respectively. Adhesion of Bifidobacterium strains to mucin preparations was markedly affected by the degree of purification. In eight of 22 strains, we observed increased adhesion to mucin preparations purified by ultracentrifugation. Moreover, in some of these eight strains, adhesion to mucin was reduced by pretreatment with sulfatase and/or sialidase, and competitively inhibited by pretreatment with PGM34 and/or HCM31 mAbs. Our results showed that some Bifidobacterium strains adhered to sulfo- and/or sialomucin and were able to recognize carbohydrate structures of the mAbs epitopes.     

EcoBalance 2018—Nexus of ideas: innovation by linking through life cycle thinking (9–12 October 2018, Tokyo,Japan)

The International Journal of Life Cycle Assessment -     

Effects of 2,6-dichlorobenzonitrile on differentiation to tracheary elements of isolated mesophyll cells of Zinnia elegans and formation of secondary cell walls

Seeds of Caesulia axillaris Roxb. displayed an absolute light requirement for germination throughout the period of dry storage at 28°C. The seeds were found to show a gradual increase in percent germination with storage time - reaching a maximum value between 8-14 months and then a sharp decline. Percent water uptake and photosensitivity were at maximum after a 5-day imbibition period in the dark in both seedlots studied. Seedlot I, which was only marginally responsive to far-red light, showed a nearly complete red-far-red reversal effect in contrast to seedlot II. The latter also displayed a considerable promotion of germination in far-red light. Interestingly, a noticeable degree of heterogeneity, besides the one observed in both seedlots with reference to red light, was found to exist in seedlot II for far-red light. Exogenous application of nitrate and ammonium, at the levels occurring in soil during seed germination/seedling emergence phase of the plant in nature, promoted a considerable proportion of high Ø-requiring seeds to germinate under irradiation conditions establishing low Ø-value. The probable ecological implication of this reponse has been discussed. Little correlation was found between the requirement for an exogenous supply of nitrate and the endogenous nitrate level in the seeds in their response to far-red light.