The central role of PhEIN2 in ethylene responses throughout plant development in petunia

The plant hormone ethylene regulates many aspects of growth and development. Loss-of-function mutations in ETHYLENE INSENSITIVE2 (EIN2) result in ethylene insensitivity in Arabidopsis, indicating an essential role of EIN2 in ethylene signaling. However, little is known about the role of EIN2 in species other than Arabidopsis. To gain a better understanding of EIN2, a petunia (Petunia x hybrida cv Mitchell Diploid [MD]) homolog of the Arabidopsis EIN2 gene (PhEIN2) was isolated, and the role of PhEIN2 was analyzed in a wide range of plant responses to ethylene, many that do not occur in Arabidopsis. PhEIN2 mRNA was present at varying levels in tissues examined, and the PhEIN2 expression decreased after ethylene treatment in petals. These results indicate that expression of PhEIN2 mRNA is spatially and temporally regulated in petunia during plant development. Transgenic petunia plants with reduced PhEIN2 expression were compared to wild-type MD and ethylene-insensitive petunia plants expressing the Arabidopsis etr1-1 gene for several physiological processes. Both PhEIN2 and etr1-1 transgenic plants exhibited significant delays in flower senescence and fruit ripening, inhibited adventitious root and seedling root hair formation, premature death, and increased hypocotyl length in seedling ethylene response assays compared to MD. Moderate or strong levels of reduction in ethylene sensitivity were achieved with expression of both etr1-1 and PhEIN2 transgenes, as measured by downstream expression of PhEIL1. These results demonstrate that PhEIN2 mediates ethylene signals in a wide range of physiological processes and also indicate the central role of EIN2 in ethylene signal transduction.     

A‐FABP—A Biomarker Associated With the Metabolic Syndrome and/or an Indicator of Weight Change?

Objective: Adipocyte fatty acid‐binding protein (A‐FABP) is a plasma biomarker recently associated with the metabolic syndrome. The aim of these studies was to investigate changes of A‐FABP during profound weight loss induced by laparoscopic adjustable gastric banding (LAGB). Methods and Procedures: In study one, 29 severely obese female subjects were examined before and 1 year after surgical treatment. A subgroup of 10 patients was investigated in 3‐month intervals. Metabolic parameters were determined using standard methods, and A‐FABP was detected using a commercially available enzyme‐linked immunosorbent assay. Results: Mean weight loss after 1 year was 24.9 kg (P < 0.001), mainly due to a decrease in fat mass. Metabolic parameters improved substantially. However, serum A‐FABP remained stable. In study two, a subgroup of 10 patients was examined quarterly to determine the time course of A‐FABP changes. Quarterly measurements of serum A‐FABP were significantly higher than baseline levels with the highest A‐FABP value after the first 3 months, where patients had highest weight loss. Discussion: Our results in study one show that A‐FABP serum levels are positively associated with body weight and fat mass. However, 1 year after pronounced weight loss A‐FABP levels remained unchanged. In study two, time course analyses revealed maximum increase of serum A‐FABP in parallel to highest weight loss, which allows to suppose that A‐FABP is not only a biomarker of the metabolic syndrome in the steady state, but also a marker of weight changes in dynamic situations.     

Chlamydia trachomatis detection in a population of asymptomatic and symptomatic women: correlation with the presence of serological markers for this infection

A study of 371 women (261 asymptomatic and 110 symptomatic subjects with clinical PID) was performed to detect the presence of Chlamydia trachomatis (C.t.) and to correlate the serological markers against this microrganism, such as antibody to chlamydial hsp60 (Ab-Chsp60) and different levels of IgG, IgM and IgA, with epidemiology, pathology, sexual habits, age, diagnostic methods in the groups of women with and without pelvic inflammatory disease (PID). We found a statistically significant difference between the asymptomatic and symptomatic women regarding the presence of C.t. (3.4% versus 20%; p<0.0001). This presence was affected by the age of women (more in the group < or =25 years old), by having sex with new partners mainly if they did not undergo an antibiotic treatment. The association of antibody Chsp60 with the presence of clinical PID was quite striking. We also found a strict correlation between the detection of Ab-Chsp60 and previous chlamydial infection as well as between Ab-Chsp60 and elevated serum chlamydial IgG or IgA levels. Due to these findings, we can say that the use of serological markers for C.t. in clinical practice may be an important tool for an early screening and diagnosis of women at high risk of chlamydial infection.     

Pioglitazone has direct effects on insulin sensitivity and intracellular lipid content in L6 skeletal muscle cells

Using L6 skeletal muscle cell line, rendered insulin resistant by incubation with triglyceride-rich lipoproteins (TGRLs), we sought to answer the question whether pioglitazone has direct effects on this cell line. Incubation of L6 cells with TGRLs led to an increase in the intramyocellular triglyceride content. Moreover, TGRLs led to a reduction in insulin-stimulated glycogen content and GSK-3 phosphorylation. All these changes induced by TGRLs could be antagonized by incubation of L6 cells with pioglitazone. The PPAR-γ antagonist GW9662 reversed the pioglitazone effects. We conclude that pioglitazone has direct insulin-sensitizing effects on the L6 skeletal muscle cell line, which are paralleled by a reduction in intramyocellular triglyceride accumulation.     

Ethylene-dependent salicylic acid regulates an expanded cell death response to a plant pathogen

The molecular events associated with susceptible plant responses to disease-causing organisms are not well understood. We have previously shown that ethylene-insensitive tomato plants infected with Xanthomonas campestris pv. vesicatoria have greatly reduced disease symptoms relative to wild-type cultivars. Here we show that salicylic acid (SA) is also an important component of the susceptible disease response. SA accumulates in infected wild-type tissues and is correlated with necrosis but does not accumulate in ethylene-insensitive plants. Exogenous feeding of SA to ethylene-deficient plants restores necrosis, indicating that reduced disease symptoms are associated with failure to accumulate SA. These results indicate a mechanism for co-ordination of phytohormone signals that together constitute a susceptible response to pathogens.     

A Practical and Stereoselective Organocatalytic Alkylation of Aldehydes with Benzodithiolylium Tetrafluoroborate

Recently, the direct substitution of allylic, benzylic, and tertiary alcohols has been achieved via S_N1‐type reactions with catalytic amounts of Brønsted or Lewis acids. When a new stereogenic center is formed most of these transformations produce the desired product as a racemate, as these reactions proceed through carbenium ions. The arsenal of activation modes available in organocatalysis can be used to set up suitable reaction conditions in which chiral nucleophiles (enamine catalysis) or chiral electrophiles (iminium catalysis, chiral counterion catalysis) can easily be generated. Recently, we have used stabilized carbenium ions, directly available or obtained from the corresponding alcohols, in new organocatalytic stereoselective S_N1‐type reactions. The commercially available carbenium ion benzodithiolylium tetrafluoroborate 1 can be used for the straightforward organocatalytic stereoselective alkylation of aldehydes. In this account we will illustrate the application of this methodology in the total synthesis of natural products and the preparation of valuable starting materials. Chirality 26:607–613, 2014. © 2014 Wiley Periodicals, Inc.     

MiR-424 and miR-155 deregulated expression in cytogenetically normal acute myeloid leukaemia: correlation with NPM1 and FLT3 mutation status

Background

Gain-of-function mutations of tyrosine kinase FLT3 are frequently found in acute myeloid leukemia (AML). This has made FLT3 an important marker for disease diagnosis and a highly attractive target for therapeutic drug development. This study is intended to generate a sensitive substrate for assays of the FLT3 enzymatic activity.

Methods

We expressed in Escherichia coli cells a glutathione S-transferase (GST) fusion protein designated GST-FLT3S, which contains a peptide sequence derived from an autophosphorylation site of FLT3. The protein was used to analyze tyrosine kinase activity of baculovirus-expressed FLT3 and crude cell extracts of bone marrow cells from AML patients. It was also employed to perform FLT3 kinase assays for FLT3 inhibitor screening.

Results

GST-FLT3S in solution or on beads was strongly phosphorylated by recombinant proteins carrying the catalytic domain of wild type FLT3 and FLT3D835 mutants, with the latter exhibiting much higher activity and efficiency. GST-FLT3S was also able to detect elevated tyrosine kinase activity in bone marrow cell extracts from AML patients. A small-scale inhibitor screening led to identification of several potent inhibitors of wild type and mutant forms of FLT3.

Conclusions

GST-FLT3S is a sensitive protein substrate for FLT3 assays. It may find applications in diagnosis of diseases related to abnormal FLT3 activity and in inhibitor screening for drug development.     

Free circulating ICAM-1 in serum and cerebrospinal fluid of HIV-1 infected patients correlate with TNF-alpha and blood-brain barrier damage

The mechanism for the initiation of blood-brain barrier damage and intrathecal inflammation in patients infected with the human immunodeficiency virus (HIV) is poorly understood. We have recently reported that tumour necrosis factor-alpha (TNF-alpha) mediates active neural inflammation and blood-brain barrier damage in HIV-1 infection. Stimulation of endothelial cells by TNF-alpha induces the expression of intercellular adhesion molecule-1 (ICAM-1), which is an important early marker of immune activation and response. We report herein for the first time the detection of high levels of free circulating ICAM-1 in serum and cerebrospinal fluid of patients with HIV-1 infection. Free circulating ICAM-1 in these patients correlated with TNF-alpha concentrations and with the degree of blood-brain barrier damage and were detected predominantly in patients with neurologic involvement. These findings have important implications for the understanding and investigation of the intrathecal inflammatory response in HIV-1 infection.     

Regulation of Ethylene-mediated Responses at the Level of the Receptor

The ethylene receptor is currently the best characterized ofplant hormone receptors. Mutant and biochemical analyses haverevealed much about the mode(s) of action and biological regulationof ethylene responses. The receptors are related to bacterialtwo-component regulators, which are histidine protein kinases.In both arabidopsis and tomato, a family of genes encodes astructurally divergent set of receptors. While one arabidopsisreceptor, ETR1, has been shown to be a functional histidinekinase, the significant structural divergence among the familymembers makes the actual mode of action of the receptors unclear.Molecular and genetic evidence indicates that quantitative alterationsin receptor levels can modify the sensitivity of a plant ortissue to ethylene. However, our current knowledge of receptorgene regulation does not address how a plant regulates differentialresponsiveness of a tissue to ethylene. Copyright 2001 Annalsof Botany Company Ethylene sensitivity, ERS, ETR, histidine kinase, LeETR, Lycopersicon esculentum