Indole-3-ethanol oxidase in Phycomyces blakesleeanus. Is indole-3-ethanol a “storage pool” for IAA?

Indole-3-ethanol (IEt) was extracted from Phycomyces blakesleeanus Bgff. and purified by TLC and HPLC. Identification was performed by mass spectrum. The HPLC-purified compound showed an UV-spectrum typical for indoles, with absorption maxima at 220 and 281 nm. The IEt content varied between 1.5 nmol (g fresh weight)⁻¹ and 5.6 nmol (g fresh weight)⁻¹. The observed variations were strongly correlated with certain developmental stages of the fungus. Furthermore, the decrease of IEt between 60 and 84 h of fungal development coincides with a high IEt oxidase activity. The product of the enzyme reaction was indole-3-acetaldehyde, which was identified by co-chromatography with an authentic standard in several TLC and HPLC systems and by chemical conversion to indole-3-acetaldoxime.     

Active Glucose Transport and Proton Pumping in Tonoplast Membrane of Zea mays L. Coleoptiles Are Inhibited by Anti-H-ATPase Antibodies

A tonoplast enriched fraction was obtained from Zea mays L. coleoptiles by isopycnic centrifugation of microsomal membranes in a sucrose step gradient. At the 18/26% interface chloride-stimulated and nitrate-inhibited proton pumping activity coincided with a Mg²⁺-ATP dependent accumulation of 3-O-methyl-d-glucose (OMG) as determined by a membrane filtration technique using ¹⁴C-labeled substrate. OMG transport showed an apparently saturable component with a K_m of 110 micromolar, and was completely inhibited by 10 micromolar carbonyl cyanide m-chlorophenylhydrazone. Polyclonal antibodies against solubilized native tonoplast H⁺-ATPase and its 62 and 72 kilodalton subunits were assayed for their ability to inhibit proton pumping and OMG accumulation. Antibodies against both the native enzyme and the putative catalytic subunit (72 kilodalton) strongly inhibited proton pumping and OMG transport whereas antibodies against the 62 kilodalton subunit had only a slight effect on both processes.     

Stimulation of pancreatic secretory process in the rat by low-molecular weight proteinase inhibitor

Oral application of a single dose of a new synthetic proteinase inhibitor Camostate (Foy-305) in male Wistar rats was carried out together with studies of in vitro amino acid incorporation followed by separation of proteins by two-dimensional gel electrophoresis. The aim of this experiment was to analyze changes produced by the inhibitor in total protein and individual enzyme biosynthesis. Administration of 100 mg/kg Foy-305 resulted in significant inhibition of total pancreatic protein synthesis, without changes in fractional rates for individual enzymes. 50 mg/kg Foy-305 induced a 10-fold elevation of cholecystokinin (CCK) levels in serum; this persisted for 3 h and led to a significant increase in the total rate of protein synthesis with peak values at 6 and 9 h (78% and 84% above control levels, respectively), returning to control by 15 h. Changes in fractional rates of synthesis occurred with a latency of 6 h and were restricted to amylase and the anionic form of trypsinogen and chymotrypsinogen. Amylase biosynthesis decreased by about 40% from control levels at 9 h to return to control levels by 15 h. Increased synthesis of trypsinogen and chymotrypsinogen was observed; this was also phasic. The results show similar enzyme-specific regulation as previously described for exogenous CCK stimulation and for the adaptation of the pancreas to diets enriched in protein. They demonstrate the effectiveness of pulsatory endogenous hormone release in the regulation of protein synthesis.     

Functional expression of dihydropyridine-insensitive calcium channels during PC12 cell differentiation by nerve growth factor (NGF), oncogenic ras,or src tyrosine kinase

1. Recombinant retroviruses were used to introduce a temperature-sensitive v-src gene and oncogenic c-Ha-ras into PC12 cells, and stable cell lines expressing these genes were established. 2. As previously reported, expression of v-src (Alema et al., 1985) or c-Ha-ras (Noda et al., 1985) in PC12 cells results in neurite outgrowth resembling that induced by NGF. We report here that v-src but not oncogenic c-Ha-ras induces a stable morphologic neuronal differentiation similar to treatment with NGF. Oncogenic c-Ha-ras-induced neurite outgrowth is not stable with long-term culture, rather the cells revert to an undifferentiated morphology with altered cell cycle kinetics. 3. The stable neuronal phenotype induced by v-src and NGF is characterized by the functional expression of dihydropyridine-insensitive calcium currents.     

ADP Is a Competitive Inhibitor of ATP-Dependent H Transport in Microsomal Membranes from Zea mays L. Coleoptiles

An anion-sensitive ATP-dependent H⁺ transport in microsomal membranes from Zea mays L. coleoptiles was partially characterized using the pH gradient-dependent decrease of unprotonated neutral red. The following criteria strongly suggest a tonoplast origin of the H⁺ transport observed: strict dependence on Cl⁻; inhibition by SO₄²⁻ and NO₃⁻; insensitivity against vanadate, molybdate, and azide; reversible inhibition by CaCl₂ (H⁺/Ca²⁺ antiport); inhibition by diethylstilbestrol. The substrate kinetics revealed simple Michaelis Menten kinetics for ATP in the presence of 1 millimolar MgCl₂ with a K_m value of 0.56 millimolar (0.38 millimolar for MgATP). AMP and c-AMP did not influence H⁺ transport significantly. However, ADP was a potent competitive inhibitor with a K_i value of 0.18 millimolar. The same inhibition type was found for membranes prepared from primary roots by the same procedure.     

Comparative effects of trichothecene mycotoxins on bovine platelet function: Acetyl T-2 toxin,a more potent inhibitor than T-2 toxin

The effects of the trichothecene mycotoxins (acetyl T-2 toxin, T-2 toxin, HT-2 toxin, palmityl T-2 toxin, diacetoxyscirpenol (DAS), deoxynivalenol (DON), and T-2 tetraol) on bovine platelet function were examined in homologous plasma stimulated with platelet activating factor (PAF). The mycotoxins inhibited platelet function with the following order of potency: acetyl T-2 toxin > palmityl T-2 toxin = DAS > HT-2 toxin = T-2 toxin. While T-2 tetraol was completely ineffective as an inhibitor, DON exhibited minimal inhibitory activity at concentrations above 10×10^?4M. The stability of the platelet aggregates formed was significantly reduced in all mycotoxin treated platelets compared to that of the untreated PAF controls. It is suggested that the increased sensitivity of PAF stimulated bovine platelets to the more lipophilic mycotoxins may be related to their more efficient partitioning into the platelet membrane compared to the more hydrophilic compounds.     

An extensive review on antifungal approaches in the treatment of mucormycosis

During the period of COVID-19, the occurrences of mucormycosis in immunocompromised patients have increased significantly. Mucormycosis (black fungus) is a rare and rapidly progressing fungal infection associated with high mortality and morbidity in India as well as globally. The causative agents for this infection are collectively called mucoromycetes which are the members of the order Mucorales. The diagnosis of the infection needs to be performed as soon as the occurrence of clinical symptoms which differs with types of Mucorales infection. Imaging techniques magnetic resonance imaging or computed tomography scan, culture testing, and microscopy are the approaches for the diagnosis. After the diagnosis of the infection is confirmed, rapid action is needed for the treatment in the form of antifungal therapy or surgery depending upon the severity of the infection. Delaying in treatment declines the chances of survival. In antifungal therapy, there are two approaches first-line therapy (monotherapy) and combination therapy. Amphotericin B ( 1 ) and isavuconazole ( 2 ) are the drugs of choice for first-line therapy in the treatment of mucormycosis. Salvage therapy with posaconazole ( 3 ) and deferasirox ( 4 ) is another approach for patients who are not responsible for any other therapy. Adjunctive therapy is also used in the treatment of mucormycosis along with first-line therapy, which involves hyperbaric oxygen and cytokine therapy. There are some drugs like VT-1161 ( 5 ) and APX001A ( 6 ), Colistin, SCH 42427, and PC1244 that are under clinical trials. Despite all these approaches, none can be 100% successful in giving results. Therefore, new medications with favorable or little side effects are required for the treatment of mucormycosis.     

Reduction of levels of nuclear-associated protein in heated cells by cycloheximide, D2O, and thermotolerance.

Hyperthermia increases levels of nuclear-associated proteins in a manner that correlates with cell killing. If the increase in nuclear-associated proteins represents a lethal lesion then treatments that protect against killing by heat should reduce and/or facilitate the recovery of levels of the proteins in heated cells. This hypothesis was tested using three heat protection treatments: cycloheximide, D2O, and thermotolerance. All three treatments reduced levels of the proteins measured immediately following hyperthermia at 43.0 or 45.5 degrees C, with the greatest reduction occurring at 43.0 degrees C. In addition to reducing the proteins, thermotolerance facilitated the recovery of the proteins to control levels following hyperthermia. Thus thermotolerance may protect cells by both reducing the initial heat damage and facilitating recovery from that damage. Cycloheximide and D2O did not facilitate recovery of nuclear-associated proteins, suggesting that their protection against cytotoxicity related to the proteins resulted solely from their reduction of increases in levels of the proteins. All three treatments have been shown to stabilize cellular proteins against thermal denaturation. The results of this study suggest that the increase in nuclear-associated proteins may result from thermally denatured proteins adhering to the nucleus and that it is the ability of cycloheximide, D2O, and thermotolerance to thermostabilize proteins that reduces the increase in levels of the proteins within heated cells.     

Primary Action of Indole-3-acetic Acid in Crown Gall Tumors: Increase of Solute Uptake

Exogenously added indole-3-acetic acid at a concentration of 100 micromolars stimulates d-glucose uptake (or 3-O-methyl-d-glucose uptake) by 25% in crown gall tumors induced on potato tuber tissue by Agrobacterium tumefaciens strain C 58. The titration of the endogenous IAA with the auxin antagonist 2-naphthaleneacetic acid at 100 micromolars reduces d-glucose uptake by about 80%. The apparent inhibition constant K_i is 21 micromolars. Other auxin antagonists like 1-naphthoxyacetic acid and 2-(p-chlorophenoxy)-2-methylpropionic acid show similar effects. The uptake of the amino acids leucine, methionine, tryptophan, lysine, and aspartic acid is also inhibited by 2-naphthaleneacetic acid to similar degrees. The auxins 1-naphthaleneacetic acid and 2-naphthoxyacetic acid at concentrations between 10 and 100 micromolars inhibit solute uptake only slightly (inhibition less than 20%). The impact of the results on the postulated role of indole-3-acetic acid as a modifier of the electrochemical proton gradient across the plasmalemma in crown gall tumor tissue is discussed.