Hexane fraction of Globimetula braunii induces mitochondria-mediated apoptosis in Plasmodium berghei-infected mice.

Background: Apoptosis is a common pathology in malaria and most antimalarial drugs induce apoptosis during chemotherapy. Globimetula braunii is an African mistletoe used for the treatment of malaria but its effect on mitochondria-mediated apoptosis is not known. Methods: Malarial infection was induced by the intraperitoneal injection of NK 65 strain Plasmodium berghei-infected erythrocytes into mice which were treated with graded doses (100-400 mg/kg) of methanol extract (ME), and fractions of n-hexane, dichloromethane, ethylacetate and methanol (HF, DF, EF and MF) for 9 days after the confirmation of parasitemia. Artequine (10 mg/kg) was used as control drug. The fraction with the highest antiplasmodial activity was used (same dose) to treat mice infected with chloroquine-resistant (ANKA) strain for 5 consecutive days after the confirmation of parasitemia. P-alaxin (10 mg/kg) was used as control drug. On the last day of the treatment, liver mitochondria were isolated and mitochondrial Permeability Transition (mPT) pore opening, mitochondrial F0F1 ATPase (mATPase) activity, lipid peroxidation (mLPO) and liver deoxyribonucleic acid (DNA) fragmentation were assessed spectrophotometrically. Caspases 3 and 9 were determined by Enzyme-Linked Immunosorbent Assay (ELISA) technique. Cytochrome c, P53, Bcl-2-associated X protein (Bax), and B-cell lymphoma-2 (Bcl2) were determined via immunohistochemistry. Phytochemical constituents of the crude methanol extract of Globimetula braunii were determined via the Gas Chromatography-Mass Spectrometry (GC-MS) analysis. Results: There was large amplitude mPT induction by malaria parasites, extract and fractions of Globimetula braunii. At 400 mg/kg, HF significantly (p < 0.01) downregulated mATPase activity, and mLPO in both (susceptible and resistant) models, caused DNA fragmentation (P < 0.0001), induced caspases activation, P53, bax and cytochrome c release but downregulated Bcl2 in both models. The GC-MS analysis of methanol extract of Globimetula braunii showed that α-amyrin is the most abundant phytochemical. Conclusion: The n-hexane fraction of Globimetula braunii induced mitochondrial-mediated apoptosis through the opening of the mitochondrial pore, fragmentation of genomic DNA, increase in the levels of P53, bax, caspase 3 and 9 activation and cytochrome c release with concomitant decrease in the level of Bcl2. α-Amyrin is a triterpene with apoptotic effects.

A distributable LC-MS/MS method for the measurement of serum thyroglobulin.

Background: Despite its clear advantages over immunoassay-based testing, the measurement of serum thyroglobulin by mass spectrometry remains limited to a handful of institutions. Slow adoption by clinical laboratories could reflect limited accessibility to existing methods that have sensitivity comparable to modern immunoassays, as well as a lack of tools for calibration and assay harmonization. Methods: We developed and validated a liquid chromatography-tandem mass spectrometry-based assay for the quantification of serum thyroglobulin. The protocol combined peptide immunoaffinity purification using a commercially available, well-characterized monoclonal antibody and mobile phase modification with dimethylsulfoxide (DMSO) for enhanced sensitivity. To facilitate harmonization with other laboratories, we developed a novel, serum-based 5-point distributable reference material (Husky Ref). Results: The assay demonstrated a lower limit of quantification of 0.15 ng/mL (<20 %CV). Mobile phase DMSO increased signal intensity of the target peptide at least 3-fold, improving quantification at low concentrations. Calibration traceable to Husky Ref enabled harmonization between laboratories in an interlaboratory study. Conclusions: Sensitive mass spectrometry-based thyroglobulin measurement can be achieved using a monoclonal antibody during peptide immunoaffinity purification and the addition of mobile phase DMSO. Laboratories interested in deploying this assay can utilize the provided standard operating procedure and freely-available Husky Ref reference material.

Recommendations for the use of the acetaminophen hepatotoxicity model for mechanistic studies and how to avoid common pitfalls.

Acetaminophen (APAP) is a widely used analgesic and antipyretic drug, which is safe at therapeutic doses but can cause severe liver injury and even liver failure after overdoses. The mouse model of APAP hepatotoxicity recapitulates closely the human pathophysiology. As a result, this clinically relevant model is frequently used to study mechanisms of drug-induced liver injury and even more so to test potential therapeutic interventions. However, the complexity of the model requires a thorough understanding of the pathophysiology to obtain valid results and mechanistic information that is translatable to the clinic. However, many studies using this model are flawed, which jeopardizes the scientific and clinical relevance. The purpose of this review is to provide a framework of the model where mechanistically sound and clinically relevant data can be obtained. The discussion provides insight into the injury mechanisms and how to study it including the critical roles of drug metabolism, mitochondrial dysfunction, necrotic cell death, autophagy and the sterile inflammatory response. In addition, the most frequently made mistakes when using this model are discussed. Thus, considering these recommendations when studying APAP hepatotoxicity will facilitate the discovery of more clinically relevant interventions.

First small-molecule PROTACs for G protein-coupled receptors: inducing α 1A-adrenergic receptor degradation.

Proteolysis targeting chimeras (PROTACs) are dual-functional hybrid molecules that can selectively recruit an E3 ubiquitin ligase to a target protein to direct the protein into the ubiquitin-proteasome system (UPS), thereby selectively reducing the target protein level by the ubiquitin-proteasome pathway. Nowadays, small-molecule PROTACs are gaining popularity as tools to degrade pathogenic protein. Herein, we present the first small-molecule PROTACs that can induce the α 1A-adrenergic receptor (α 1A-AR) degradation, which is also the first small-molecule PROTACs for G protein-coupled receptors (GPCRs) to our knowledge. These degradation inducers were developed through conjugation of known α 1-adrenergic receptors (α 1-ARs) inhibitor prazosin and cereblon (CRBN) ligand pomalidomide through the different linkers. The representative compound 9c is proved to inhibit the proliferation of PC-3 cells and result in tumor growth regression, which highlighted the potential of our study as a new therapeutic strategy for prostate cancer.

Identification of a novel PAK1 inhibitor to treat pancreatic cancer.

Pancreatic cancer is one of the most aggressive cancers with poor prognosis and a low 5-year survival rate. The family of P21-activated kinases (PAKs) appears to modulate many signaling pathways that contribute to pancreatic carcinogenesis. In this work, we demonstrated that PAK1 is a critical regulator in pancreatic cancer cell growth. PAK1-targeted inhibition is therefore a new potential therapeutic strategy for pancreatic cancer. Our small molecule screening identified a relatively specific PAK1-targeted inhibitor, CP734. Pharmacological and biochemical studies indicated that CP734 targets residue V342 of PAK1 to inhibit its ATPase activity. Further in vitro and in vivo studies elucidated that CP734 suppresses pancreatic tumor growth through depleting PAK1 kinase activity and its downstream signaling pathways. Little toxicity of CP734 was observed in murine models. Combined with gemcitabine or 5-fluorouracil, CP734 also showed synergistic effects on the anti-proliferation of pancreatic cancer cells. All these favorable results indicated that CP734 is a new potential therapeutic candidate for pancreatic cancer.

Novel radioligands for imaging sigma-1 receptor in brain using positron emission tomography (PET).

The sigma-1 receptor (σ 1R) is a unique intracellular protein. σ 1R plays a major role in various pathological conditions in the central nervous system (CNS), implicated in several neuropsychiatric disorders. Imaging of σ 1R in the brain using positron emission tomography (PET) could serve as a noninvasively tool for enhancing the understanding of the disease's pathophysiology. Moreover, σ 1R PET tracers can be used for target validation and quantification in diagnosis. Herein, we describe the radiosynthesis, in vivo PET/CT imaging of novel σ 1R 11C-labeled radioligands based on 6-hydroxypyridazinone, [11C]HCC0923 and [11C]HCC0929. Two radioligands have high affinities to σ 1R, with good selectivity. In mice PET/CT imaging, both radioligands showed appropriate kinetics and distributions. Additionally, the specific interactions of two radioligands were reduced by compounds 13 and 15 (self-blocking). Of the two, [11C]HCC0929 was further investigated in positive ligands blocking studies, using classic σ 1R agonist SA 4503 and σ 1R antagonist PD 144418. Both σ 1R ligands could extensively decreased the uptake of [11C]HCC0929 in mice brain. Besides, the biodistribution of major brain regions and organs of mice were determined in vivo. These studies demonstrated that two radioligands, especially [11C]HCC0929, possessed ideal imaging properties and might be valuable tools for non-invasive quantification of σ 1R in brain.

Orthogonal assays for the identification of inhibitors of the single-stranded nucleic acid binding protein YB-1.

We have previously shown that high expression of the nucleic acid binding factor YB-1 is strongly associated with poor prognosis in a variety of cancer types. The 3-dimensional protein structure of YB-1 has yet to be determined and its role in transcriptional regulation remains elusive. Drug targeting of transcription factors is often thought to be difficult and there are very few published high-throughput screening approaches. YB-1 predominantly binds to single-stranded nucleic acids, adding further difficulty to drug discovery. Therefore, we have developed two novel screening assays to detect compounds that interfere with the transcriptional activation properties of YB-1, both of which may be generalizable to screen for inhibitors of other nucleic acid binding molecules. The first approach is a cell-based luciferase reporter gene assay that measures the level of activation of a fragment of the E2F1 promoter by YB-1. The second approach is a novel application of the AlphaScreen system, to detect interference of YB-1 interaction with a single-stranded DNA binding site. These complementary assays examine YB-1 binding to two discrete nucleic acid sequences using two different luminescent signal outputs and were employed sequentially to screen 7360 small molecule compounds leading to the identification of three putative YB-1 inhibitors.

Novel fluorescent probes of 10-hydroxyevodiamine: autophagy and apoptosis-inducing anticancer mechanisms.

Natural product evodiamine and its derivatives represent a promising class of multi-target antitumor agents. However, the clinical development of these compounds has been hampered by a poor understanding of their antitumor mechanisms. To tackle this obstacle, herein, novel fluorescent probes were designed to elucidate the antitumor mode of action of 10-hydroxyevodiamine. This compound was proven to be distributed in the mitochondria and lysosomes and to act by autophagy and apoptosis mechanisms.

Polyphenolic extract from extra virgin olive oil inhibits the inflammatory response in IL-1ß-activated synovial fibroblasts.

The polyphenolic extract (PE) from extra virgin olive oil (EVOO) has been shown to possess important anti-inflammatory and joint protective properties in murine models of rheumatoid arthritis (RA). This study was designed to evaluate the effects of PE on IL-1ß-activated human synovial fibroblasts SW982 cell line. PE from EVOO treatment inhibited IL-1ß-induced matrix metalloproteases (P<0·001), TNF-α and IL-6 production (P<0·001). Similarly, IL-1ß-induced cyclo-oxygenase-2 and microsomal PGE synthase-1 up-regulations were down-regulated by PE (P<0·001). Moreover, IL-1ß-induced mitogen-activated protein kinase (MAPK) phosphorylation and NF-κB activation were ameliorated by PE (P<0·001). These results suggest that PE from EVOO reduces the production of proinflammatory mediators in human synovial fibroblasts; particularly, these protective effects could be related to the inhibition of MAPK and NF-κB signalling pathways. Taken together, PE from EVOO probably could provide an attractive complement in management of diseases associated with over-activation of synovial fibroblasts, such as RA.

Orthogonal assays for the identification of inhibitors of the single-stranded nucleic acid binding protein YB-1

We have previously shown that high expression of the nucleic acid binding factor YB-1 is strongly associated with poor prognosis in a variety of cancer types. The 3-dimensional protein structure of YB-1 has yet to be determined and its role in transcriptional regulation remains elusive. Drug targeting of transcription factors is often thought to be difficult and there are very few published high-throughput screening approaches. YB-1 predominantly binds to single-stranded nucleic acids, adding further difficulty to drug discovery. Therefore, we have developed two novel screening assays to detect compounds that interfere with the transcriptional activation properties of YB-1, both of which may be generalizable to screen for inhibitors of other nucleic acid binding molecules. The first approach is a cell-based luciferase reporter gene assay that measures the level of activation of a fragment of the promoter by YB-1. The second approach is a novel application of the AlphaScreen system, to detect interference of YB-1 interaction with a single-stranded DNA binding site. These complementary assays examine YB-1 binding to two discrete nucleic acid sequences using two different luminescent signal outputs and were employed sequentially to screen 7360 small molecule compounds leading to the identification of three putative YB-1 inhibitors.

7-Hydroxymatairesinol improves body weight, fat and sugar metabolism in C57BJ/6 mice on a high-fat diet.

7-Hydroxymatairesinol (7-HMR) is a plant lignan abundant in various concentrations in plant foods. The objective of this study was to test HMRLignan™, a purified form of 7-HMR, and the corresponding Picea abies extract (total extract P. abies; TEP) as dietary supplements on a background of a high-fat diet (HFD)-induced metabolic syndrome in mice and in the 3T3-L1 adipogenesis model. Mice, 3 weeks old, were fed a HFD for 60 d. Subgroups were treated with 3 mg/kg body weight 7-HMR (HMRLignan™) or 10 mg/kg body weight TEP by oral administration. 7-HMR and TEP limited the increase in body weight (-11 and -13 %) and fat mass (-11 and -18 %) in the HFD-fed mice. Epididymal adipocytes were 19 and -12 % smaller and the liver was less steatotic (-62 and -65 %). Serum lipids decreased in TEP-treated mice (-11 % cholesterol, -23 % LDL and -15 % TAG) and sugar metabolism was ameliorated by both lignan preparations, as shown by a more than 70 % decrease in insulin secretion and insulin resistance. The expression of several metabolic genes was modulated by the HFD with an effect that was reversed by lignan. In 3T3-L1 cells, the 7-HMR metabolites enterolactone (ENL) and enterodiol (END) showed a 40 % inhibition of cell differentiation accompanied by the inhibited expression of the adipogenic genes PPARγ, C/EBPα and aP2. Furthermore, END and ENL caused a 10 % reduction in TAG uptake in HEPA 1-6 hepatoma cells. In conclusion, 7-HMR and TEP reduce metabolic imbalances typical of the metabolic syndrome and obesity in male mice, whereas their metabolites inhibit adipogenesis and lipid uptake in vitro.

131I-Evans blue: evaluation of necrosis targeting property and preliminary assessment of the mechanism in animal models.

Necrosis is a form of cell death, which is related to various serious diseases such as cardiovascular disease, cancer, and neurodegeneration. Necrosis-avid agents (NAAs) selectively accumulated in the necrotic tissues can be used for imaging and/or therapy of related diseases. The aim of this study was to preliminarily investigate necrosis avidity of 131I-evans blue (131I-EB) and its mechanism. The biodistribution of 131I-EB at 24 h after intravenous administration showed that the radioactivity ratio of necrotic to viable tissue was 3.41 in the liver and 11.82 in the muscle as determined by γ counting in model rats. Autoradiography and histological staining displayed preferential uptake of 131I-EB in necrotic tissues. In vitro nuclear extracts from necrotic cells exhibited 82.3% of the uptake in nuclei at 15 min, as well as 79.2% of the uptake at 2 h after 131I-EB incubation. The DNA binding study demonstrated that evans blue (EB) has strong binding affinity with calf-thymus DNA (CT-DNA) (Ksv=5.08×105 L/(mol/L)). Furthermore, the accumulation of 131I-EB in necrotic muscle was efficiently blocked by an excess amount of unlabeled EB. In conclusion, 131I-EB can not only detect necrosis by binding the DNA released from necrotic cells, but also image necrotic tissues generated from the disease clinically.

Effects of endosulfan isomers on cytokine and nitric oxide production by differentially activated RAW 264.7 cells.

Endosulfan is an organochlorine insecticide comprised of two isomers: endosulfan-α and endosulfan-ß. Endosulfan exposure has been shown to elevate some inflammatory factors, such as nitric oxide (NO) and tumor necrosis factor (TNF), in animals or cultures of animal cells. Because the two endosulfan isomers can vary in their biological activities, the goal of this study was to determine if individual endosulfan isomers differentially impact production of NO or TNF by the mouse macrophage cell RAW 264.7 at non-cytotoxic levels. We found elevated TNF with exposure to endosulfan-α (not endosulfan-ß), but only at concentrations that were cytotoxic (≥100 µM), whereas neither endosulfan isomer altered baseline levels of NO at any concentration up to 300 µM. In interferon (IFN)-γ-activated cultures, NO levels were significantly suppressed by either endosulfan isomer at 10 µM (the lowest concentration examined), whereas only endosulfan-ß significantly lowered TNF levels at non-cytotoxic concentrations. In lipopolysaccharide (LPS)-activated cultures, both endosulfan isomers significantly reduced NO, but not TNF, at non-cytotoxic concentrations. These results suggest that the endosulfan isomers have some capacity to alter inflammatory responses differentially, particularly with IFN-γ stimulation.

The Annona muricata leaf ethanol extract affects mobility and reproduction in mutant strain NB327 Caenorhabditis elegans.

The C. elegans NB327 mutant strain is characterized for the knockdown of the dic-1 gene. The dic-1 gene is homologous to the dice-1 gene in humans, encoding the protein DICE-1 as a tumor suppressor. Absence or under-regulation of the dice-1 gene can be reflected in lung and prostate cancer [17], [18]. This study evaluated the effect of EEAML on the C. elegans NB327 mutant strain. Phenotypic aspects such as morphology, body length, locomotion, and reproductive behaviour were analyzed. It is important to emphasize that the strain presents a phenotype characteristic with respect to egg laying and hatching. Reported studies showed that Annona muricata extract and its active components evidence anti-cancer and anti-tumor effects, through experimentation in vivo and in vitro models. However, neurotoxicity has been reported as a side effect. The results showed that the mutant strain NB327 was exposed to EEAML (5 mg/ml) concentration, it showed a significant decrease in average locomotion, resulting in 13 undulations in 30 s. This contrasts with the control strain's 17.5 undulations in 30 s. Similarly, the number of progenies was reduced from 188 progenies (control strain) to 114 and 92 progenies at the dose of (1 mg/ml and 5 mg/m) EEAML. The results of this study suggest that EEAML has a possible neurotoxic effect in concentrations equal to or greater than 5 mg/ml. Also, it does not have positive effects on the mutant strain of Caenorhabditis elegans NB327 phenotype.

Vitamin C acts as a hepatoprotectant in carbofuran treated rat liver slices in vitro.

Carbamates, most commonly used pesticides in agricultural practices, have been reported to produce free radicals causing deleterious effects in animals. The present study was designed to assess the carbofuran induced oxidative stress in rat liver slices in vitro and also to evaluate protective role of vitamin C by incubating them in Krebs-Ringer HEPES Buffer (KRHB) containing incubation media (Williams medium E (WME) supplemented with glucose and antibiotics) with different concentrations of carbofuran. The results demonstrated that carbofuran caused significant increase in lipid peroxidation and inhibition in the activity of hepatic superoxide dismutase (SOD) in concentration dependent manner. The data with incubation medium reflected that carbofuran at lowest concentration caused an increase in SOD activity followed by its inhibition at higher concentration. Carbofuran treatment caused inhibition in the activity of catalase in liver slices and WME incubation medium. Pre-incubation of liver slices and the WME media with vitamin C restored the values of biochemical indices tested. The results indicated that carbofuran might induce oxidative stress in hepatocytes. The pretreatment with vitamin C may offer hepatoprotection from toxicity of pesticide at low concentration only.

Qingfei Xiaoyan Wan, a traditional Chinese medicine formula, ameliorates Pseudomonas aeruginosa-induced acute lung inflammation by regulation of PI3K/AKT and Ras/MAPK pathways.

Gram-negative pathogen-induced nosocomial infections and resistance are a most serious menace to global public health. Qingfei Xiaoyan Wan (QF), a traditional Chinese medicine (TCM) formula, has been used clinically in China for the treatment of upper respiratory tract infections, acute or chronic bronchitis and pulmonary infection. In this study, the effects of QF on Pseudomonas aeruginosa-induced acute pneumonia in mice were evaluated. The mechanisms by which four typical anti-inflammatory ingredients from QF, arctigenin (ATG), cholic acid (CLA), chlorogenic acid (CGA) and sinapic acid (SPA), regulate anti-inflammatory signaling pathways and related targets were investigated using molecular biology and molecular docking techniques. The results showed that pretreatment with QF significantly inhibits the release of cytokines (TNF-α and IL-6) and chemokines (IL-8 and RANTES), reduces leukocytes recruitment into inflamed tissues and ameliorates pulmonary edema and necrosis. In addition, ATG was identified as the primary anti-inflammatory agent with action on the PI3K/AKT and Ras/MAPK pathways. CLA and CGA enhanced the actions of ATG and exhibited synergistic NF-κB inactivation effects possibly via the Ras/MAPK signaling pathway. Moreover, CLA is speculated to target FGFR and MEK firstly. Overall, QF regulated the PI3K/AKT and Ras/MAPK pathways to inhibit pathogenic bacterial infections effectively.

Qingfei Xiaoyan Wan, a traditional Chinese medicine formula, ameliorates Pseudomonas aeruginosa-induced acute lung inflammation by regulation of PI3K/AKT and Ras/MAPK pathways

Gram-negative pathogen-induced nosocomial infections and resistance are a most serious menace to global public health. Qingfei Xiaoyan Wan (QF), a traditional Chinese medicine (TCM) formula, has been used clinically in China for the treatment of upper respiratory tract infections, acute or chronic bronchitis and pulmonary infection. In this study, the effects of QF on Pseudomonas aeruginosa-induced acute pneumonia in mice were evaluated. The mechanisms by which four typical anti-inflammatory ingredients from QF, arctigenin (ATG), cholic acid (CLA), chlorogenic acid (CGA) and sinapic acid (SPA), regulate anti-inflammatory signaling pathways and related targets were investigated using molecular biology and molecular docking techniques. The results showed that pretreatment with QF significantly inhibits the release of cytokines (TNF-α and IL-6) and chemokines (IL-8 and RANTES), reduces leukocytes recruitment into inflamed tissues and ameliorates pulmonary edema and necrosis. In addition, ATG was identified as the primary anti-inflammatory agent with action on the PI3K/AKT and Ras/MAPK pathways. CLA and CGA enhanced the actions of ATG and exhibited synergistic NF-κB inactivation effects possibly via the Ras/MAPK signaling pathway. Moreover, CLA is speculated to target FGFR and MEK firstly. Overall, QF regulated the PI3K/AKT and Ras/MAPK pathways to inhibit pathogenic bacterial infections effectively.

The antitumor effect of tanshinone IIA on anti-proliferation and decreasing VEGF/VEGFR2 expression on the human non-small cell lung cancer A549 cell line.

The effects of tanshinone IIA on the proliferation of the human non-small cell lung cancer cell line A549 and its possible mechanism on the VEGF/VEGFR signal pathway were investigated. The exploration of the interaction between tanshinone IIA and its target proteins provides a feasible platform for studying the anticancer mechanism of active components of herbs. The CCK-8 assay was used to evaluate the proliferative activity of A549 cells treated with tanshinone IIA (2.5-80 µmol/L) for 24, 48 and 72 h, respectively. Flow cytometry was used for the detection of cell apoptosis and cell cycle perturbation. VEGF and VEGFR2 expression were studied by Western blotting. The binding mode of tanshinone IIA within the crystal structure of the VEGFR2 protein was evaluated with molecular docking analysis by use of the CDOCKER algorithm in Discovery Studio 2.1. The CCK-8 results showed that tanshinone IIA can significantly inhibit A549 cell proliferation in a dose- and time-dependent manner. Flow cytometry results showed that the apoptosis rate of tested group was higher than the vehicle control, and tanshinone IIA-treated cells accumulated at the S phase, which was higher than the vehicle control. Furthermore, the expression of VEGF and VEGFR2 was decreased in Western blot. Finally, molecular docking analysis revealed that tanshinone IIA could be stably docked into the kinase domain of VEGFR2 protein with its unique modes to form H-bonds with Cys917 and π-π stacking interactions with Val848. In conclusion, tanshinone IIA may suppress A549 proliferation, induce apoptosis and cell cycle arrest at the S phase. This drug may suppress angiogenesis by targeting the protein kinase domains of VEGF/VEGFR2.

Quantitative analysis of microtubule orientation in interdigitated leaf pavement cells.

Leaf pavement cells are shaped like a jigsaw puzzle in most dicotyledon species. Molecular genetic studies have identified several genes required for pavement cells morphogenesis and proposed that microtubules play crucial roles in the interdigitation of pavement cells. In this study, we performed quantitative analysis of cortical microtubule orientation in leaf pavement cells in Arabidopsis thaliana. We captured confocal images of cortical microtubules in cotyledon leaf epidermis expressing GFP-tubulinß and quantitatively evaluated the microtubule orientations relative to the pavement cell growth axis using original image processing techniques. Our results showed that microtubules kept parallel orientations to the growth axis during pavement cell growth. In addition, we showed that immersion treatment of seed cotyledons in solutions containing tubulin polymerization and depolymerization inhibitors decreased pavement cell complexity. Treatment with oryzalin and colchicine inhibited the symmetric division of guard mother cells.

A thermostable archaeal S-adenosylmethionine synthetase: a promising tool to improve the synthesis of adenosylmethionine analogs of biotechnological interest.

The naturally and widely occurring sulfonium compound, S-adenosylmethionine (AdoMet), one of nature's most versatile molecules, is biosynthesized from methionine and ATP by AdoMet synthetase or methionine adenosyltransferase (MAT) in a 2-step reaction in which the energy-rich sulfonium compound is formed by dephosphorylation of ATP. All living cells, with the only exception of some parasites and infectious agents, express MAT.