The oxylipin signal jasmonic acid is activated by an enzyme that conjugates it to isoleucine in Arabidopsis

Despite its importance in a variety of plant defense responses, our understanding of how jasmonic acid (JA) functions at the biochemical level is limited. Several amino acid conjugates of JA were tested for their ability to complement the JA-insensitive Arabidopsis thaliana mutant jar1-1. Unlike free JA, JA-Ile inhibited root growth in jar1-1 to the same extent as in the wild type, whereas JA-Val, JA-Leu, and JA-Phe were ineffective inhibitors in both genotypes. Thin-layer chromatography and gas chromatography-mass spectrometry (GC-MS) analysis of products produced in vitro by recombinant JAR1 demonstrated that this enzyme forms JA-amido conjugates with several amino acids, including JA-Ile. JA-Val, -Leu, -Ile, and -Phe were each quantified in Arabidopsis seedlings by GC-MS. JA-Ile was found at 29.6 pmole g(-1) fresh weight (FW) in the wild type but was more than sevenfold lower in two jar1 alleles. JA-Leu, -Val, and -Phe were present at only low levels in both genotypes. Expression of wild-type JAR1 in transgenic jar1-1 plants restored sensitivity to JA and elevated JA-Ile to the same level as in the wild type. The ethylene precursor 1-aminocyclopropane-1-carboxylic acid (ACC) conjugated to JA was also found in plant tissue at 18.4 pmole g(-1) FW. JA-ACC was determined not be an effective jasmonate root inhibitor, and surprisingly, was twofold higher in the mutants than in the wild type. This suggests that another JA-conjugating enzyme(s) is present in Arabidopsis. Synthesis of JA-ACC might provide a mechanism to coregulate the availability of JA and ACC for conversion to the active hormones JA-Ile and ethylene, respectively. We conclude that JAR1 is a JA-amino synthetase that is required to activate JA for optimal signaling in Arabidopsis. Plant hormone activation by conjugation to amino acids and the enzymes involved in their formation were previously unknown.     

Urinary 8-isoprostaglandin F(2alpha) level in Behçet's disease

Although its etiology remains unknown, the increased production of reactive oxygen species in Beh?et's disease (BD) have been reported. Furthermore, it has been suggested that vascular and endothelial tissue damage seen in BD is related to elevated reactive oxygen species generated by activated neutrophils from BD patients. To investigate the formation of lipid peroxidation in BD patients in vivo, urinary level of 8-isoprostaglandin F(2alpha) was quantitated by enzyme immunoassay after solid phase extraction in different clinical forms of BD patients. There was no difference in urinary level of 8-isoprostaglandin F(2alpha) between BD patient and healthy control group. There was also no difference in urinary levels of 8-isoprostaglandin F(2alpha) in subgroup analyses of BD patients, i.e. in mucocutaneous and vascular type BD patients; active and inactive BD patients. Contrary to the findings in literature, we found no difference in urinary level of 8-isoprostaglandin F(2alpha) between patients with systemic lupus erythematosus and healthy control group. These findings show no increase in lipid peroxidation despite the augmented formation of reactive oxygen species in BD patients. It may be interesting to assess formation of urinary level of 8-isoprostaglandin F(2alpha) in BD patients who do not take any medication.     

Jasmonate response locus JAR1 and several related Arabidopsis genes encode enzymes of the firefly luciferase superfamily that show activity on jasmonic,salicylic, and indole-3-acetic acids in an assay for adenylation

Jasmonic acid (JA) and related cyclopentanones are critical plant signaling molecules, but their mode of action at the molecular level is unclear. A map-based approach was used to identify the defective gene in the Arabidopsis JA response mutant jar1-1. JAR1 is 1 of 19 closely related Arabidopsis genes that are similar to the auxin-induced soybean GH3 gene. Analysis of fold predictions for this protein family suggested that JAR1 might belong to the acyl adenylate-forming firefly luciferase superfamily. These enzymes activate the carboxyl groups of a variety of substrates for their subsequent biochemical modification. An ATP-PPi isotope exchange assay was used to demonstrate adenylation activity in a glutathione S-transferase-JAR1 fusion protein. Activity was specific for JA, suggesting that covalent modification of JA is important for its function. Six other Arabidopsis genes were specifically active on indole-3-acetic acid (IAA), and one was active on both IAA and salicylic acid. These findings suggest that the JAR1 gene family is involved in multiple important plant signaling pathways.     

Biofilm formation in an ice cream plant

The sites of biofilm formation in an ice cream plant were investigated by sampling both the production line and the environment. Experiments were carried out twice within a 20-day period. First, stainless steel coupons were fixed to surfaces adjacent to food contact surfaces, the floor drains and the doormat. They were taken for the analysis of biofilm at three different production stages. Then, biofilm forming bacteria were␣enumerated and also presence of Listeria monocytogenes was monitored. Biofilm forming isolates were selected on the basis of colony morphology and Gram’s reaction; Gram negative cocci and rod, Gram positive cocci and spore forming isolates were identified. Most of the biofilm formations were seen on the conveyor belt of a packaging machine 8 h after the beginning of the production, 6.5 × 10³ cfu cm⁻². Most of the Gram negative bacteria identified belong to Enterobacteriaceae family such as Proteus, Enterobacter, Citrobacter, Shigella, Escherichia, Edwardsiella. The other Gram negative microflora included Aeromonas, Plesiomonas, Moraxella, Pseudomonas or Alcaligenes spp. were also isolated. Gram positive microflora of the ice cream plant included Staphyloccus, Bacillus, Listeria and lactic acid bacteria such as Streptococcus, Leuconostoc or Pediococcus spp. The results from this study highlighted the problems of spread of pathogens like Listeria and Shigella and spoilage bacteria. In the development of cleaning and disinfection procedures in ice cream plants, an awareness of these biofilm-forming bacteria is essential for the ice cream plants.     

Metformin reverses the effects of high glucose on human dermal fibroblasts of aged skin via downregulating RELA/p65 expression

Journal of Physiology and Biochemistry - Metformin has been successfully used as an anti-aging agent but exact molecular mechanisms of metformin in anti-aging remain unknown. Hyperglycemia during...     

4-Phenylbutyric Acid Plus Valproic Acid Exhibits the Therapeutic and Neuroprotective Effects in Acute Seizures Induced by Pentylenetetrazole

Neurochemical Research - Endoplasmic reticulum (ER) stress and apoptosis are implicated in the pathogenesis of epilepsy. Here we examine the effects of valproic acid (VA) plus 4-phenylbutyric acid...     

Hemin-dependent induction and internalization of CD38 in K562 cells

The cell surface antigen, CD38, is a bifunctional ecto-enzyme, which is predominantly expressed on hematopoietic cells during differentiation. In the present study, it is shown that hemin treatment of K562 cells gives rise to induction of enzymatic activities inherent to CD38. GDP-ribosyl cyclase activity, an indicator of CD38, increased initially in response to hemin in a time-dependent manner, reached a maximum level on the 5th day and, thereafter, declined sharply to the initial level. The increase in NAD(+) glycohydrolase and ADP-ribose uptake activities followed a similar time course. However, the decline in the latter activities after the 5th day of induction appeared to be rather slow in contrast to GDP-ribosyl cyclase activity. The time course of these changes was well correlated with the FACScan findings obtained by use of anti-CD38 monoclonal antibody. SDS-PAGE and Western blot analyses by use of the monoclonal antibody OKT10 revealed a transient hemin-dependent appearance of a 43 kDa membrane protein with maximum signal intensity on the first 4 days of incubation. There was subsequently a gradual decrease on the 5th day, concomitant with a reciprocal increase in activity of the internalized protein fraction. The results together indicated that hemin-induced expression of CD38 was followed by its down-regulation.