Induction of Erythroid Differentiation in Human Erythroleukemia Cells by Depletion of Malic Enzyme 2

Malic enzyme 2 (ME2) is a mitochondrial enzyme that catalyzes the conversion of malate to pyruvate and CO2 and uses NAD as a cofactor. Higher expression of this enzyme correlates with the degree of cell de-differentiation. We found that ME2 is expressed in K562 erythroleukemia cells, in which a number of agents have been found to induce differentiation either along the erythroid or the myeloid lineage. We found that knockdown of ME2 led to diminished proliferation of tumor cells and increased apoptosis in vitro. These findings were accompanied by differentiation of K562 cells along the erythroid lineage, as confirmed by staining for glycophorin A and hemoglobin production. ME2 knockdown also totally abolished growth of K562 cells in nude mice. Increased ROS levels, likely reflecting increased mitochondrial production, and a decreased NADPH/NADP+ ratio were noted but use of a free radical scavenger to decrease inhibition of ROS levels did not reverse the differentiation or apoptotic phenotype, suggesting that ROS production is not causally involved in the resultant phenotype. As might be expected, depletion of ME2 induced an increase in the NAD+/NADH ratio and ATP levels fell significantly. Inhibition of the malate-aspartate shuttle was insufficient to induce K562 differentiation. We also examined several intracellular signaling pathways and expression of transcription factors and intermediate filament proteins whose expression is known to be modulated during erythroid differentiation in K562 cells. We found that silencing of ME2 leads to phospho-ERK1/2 inhibition, phospho-AKT activation, increased GATA-1 expression and diminished vimentin expression. Metabolomic analysis, conducted to gain insight into intermediary metabolic pathways that ME2 knockdown might affect, showed that ME2 depletion resulted in high orotate levels, suggesting potential impairment of pyrimidine metabolism. Collectively our data point to ME2 as a potentially novel metabolic target for leukemia therapy

Thermal and structural response of in situ prepared biobased poly(ethylene 2,5-furan dicarboxylate) nanocomposites

Poly(ethylene 2,5-furan dicarboxylate) (PEF) is considered the biobased counterpart of the fossil based poly(ethylene terephthalate) for food packaging. In this research, PEF nanocomposites containing 2.5 wt% neat multi walled carbon nanotubes (MWCNTs), or functionalized MWCNTs or graphene oxide (GO), were in situ prepared by applying the melt polycondensation method. The nanocomposites showed faster crystallization rates compared to the pristine material as proved by both differential scanning calorimetry (DSC) and polarized light microscopy (PLM). The latter evidenced an increased nucleation density in nanocomposites, due to the nucleating efficiency of the fillers, resulting in smaller spherulite size. However, a slightly reduced thermal stability was revealed for the nanocomposites by thermog-ravimetric analysis (TGA), especially in the case of GO-containing samples. The solid structure of the materials was studied by performing real time X-ray diffraction (XRD) measurements. In neat PEF, beta-crystals were observed in the solvent treated sample, while alpha-crystals were formed on cooling from the melt or cold-crystallization. On the contrary, in the XRD patterns of the nanocomposites only peaks associated with the alpha-crystal phase were found. Last, but not least, the effect of recrystallization on the thermal behavior was evaluated by means of modulated temperature DSC (MDSC). (C) 2016 Elsevier Ltd. All rights reserved

Trapping of Intermediates with Substrate Analog HBOCaA in the Polymerizations Catalyzer by Class III Polyhydroxybutyrate (PHB) Synthase from Allochromatium Vinosum

Polyhydroxybutyrate (PHB) synthases (PhaCs) catalyze the formation of biodegradable PHB polymers that are considered as an ideal alternative to petroleum-based plastics. To provide strong evidence for the preferred mechanistic model involving covalent and noncovalent intermediates, a substrate analog HBOCoA was synthesized chemoenzymatically. Substitution of sulfur in the native substrate HBCoA with an oxygen in HBOCoA enabled detection of (HB)nOCoA (n = 2–6) intermediates when the polymerization was catalyzed by wild-type (wt-)PhaECAv at 5.84 hr−1. This extremely slow rate is due to thermodynamically unfavorable steps that involve formation of enzyme-bound PHB species (thioesters) from corresponding CoA oxoesters. Synthesized standards (HB)nOCoA (n = 2–3) were found to undergo both reacylation and hydrolysis catalyzed by the synthase. Distribution of the hydrolysis products highlights the importance of the penultimate ester group as previously suggested. Importantly, the reaction between primed synthase [3H]-sT-PhaECAv and HBOCoA yielded [3H]-sTet-O-CoA at a rate constant faster than 17.4 s−1, which represents the first example that a substrate analog undergoes PHB chain elongation at a rate close to that of the native substrate (65.0 s−1). Therefore, for the first time with a wt-synthase, strong evidence was obtained to support our favored PHB chain elongation model

Loss of Mitogen-Activated Protein Kinase Kinase Kinase 4 (MAP3K4) Reveals a Requirement for MAPK Signalling in Mouse Sex Determination

The boygirl (byg) mouse mutant reveals that MAP3K4-mediated signaling is necessary for normal SRY expression and testis specification in the developing mouse gonad

Evaluaci?n de hongos entomopat?genos para el control microbiano de Bemisia tabaci (Hom?ptera: aleyrodidae)

Tesis (M. Sc) -- CATIE, Turrialba (Costa Rica), 1995Se probaron en laboratorio 29 aislamientos de Beauveria sp., uno de B. brongniartii, 10 de Metarhizium anisopliae, uno de M. flavoviride uno de Paecilomyces fumosoroseus y uno de Paecilomyces sp. provenientes de las colecciones del CATIE (Costa Rica y Nicaragua) y del USDA (Departamento de Agricultura de Estados Unidos). La metodolog?a de bioensayos consisti? en cortar discos de hoja de frijol con ninfas de cuarto estadio de B. tabaci. Estos discos se sumergieron en una suspensi?n de 1*10 exponente 7 conidios/ml y se evalu? la mortalidad causada por cada aislado. La mayor?a de aislamientos no tuvo ning?n efecto marcado sobre las ninfas de B. tabaci. Algunos aislamientos de B. bassiana y P. fumosoroseus fueron diferentes al testigo, pero el porcentaje de mortalidad no sobrepas? el 47 por ciento. Cinco aislamientos de M. anisopliae fueron muy virulentos con hasta 97 por ciento de mortalidad