A tRNA(Glu) that uncouples protein and tetrapyrrole biosynthesis

Glu-tRNA is either bound to elongation factor Tu to enter protein synthesis or is reduced by glutamyl-tRNA reductase (GluTR) in the first step of tetrapyrrole biosynthesis in most bacteria, archaea and in chloroplasts. Acidithiobacillus ferrooxidans, a bacterium that synthesizes a vast amount of heme, contains three genes encoding tRNA(Glu). All tRNA(Glu) species are substrates in vitro of GluRS1 from A. ferrooxidans.Glu-tRNA(3)(Glu), that fulfills the requirements for protein synthesis, is not substrate of GluTR. Therefore, aminoacylation of tRNA(3)(Glu) might contribute to ensure protein synthesis upon high heme demand by an uncoupling of protein and heme biosynthesis.     

12-O-tetradecanoylphorbol-13-acetate may both potentiate and decrease the generation of apoptosis by the antileukemic agent arsenic trioxide in human promonocytic cells. Regulation by extracellular signal-regulated protein kinases and glutathione

Arsenic trioxide (As(2)O(3)) caused apoptosis in U-937 human promonocytic cells. This effect was potentiated by the simultaneous addition of the glutathione (GSH) synthesis inhibitor DL-buthionine-(R,S)-sulfoximine or the protein kinase C activators 12-O-tetradecanoylphorbol-13-acetate (TPA) and bryostatin 1. In addition TPA decreased the intracellular GSH content, caused ERK activation, and potentiated the As(2)O(3)-provoked activation of p38 and JNK. The addition of N-acetyl-L-cysteine, the PKC inhibitor GF109203X, and the MEK/ERK inhibitors PD98059 and U0126 attenuated both apoptosis induction and GSH decrease, whereas the p38 inhibitor SB203580 and the JNK inhibitor SP600125 were ineffective. TPA also potentiated ERK activation and GSH depletion when added simultaneously to cadmium chloride (CdCl(2)) and doxorubicin. However, TPA only enhanced apoptosis in the case of CdCl(2), which is a GSH-sensitive agent, whereas it reduced the toxicity of doxorubicin and other DNA-specific drugs. Finally, preincubation for 14-24 h with TPA did not potentiate but, instead, attenuated the As(2)O(3)- and CdCl(2)-provoked apoptosis. The same result was obtained by preincubation with bryostatin 1 and other differentiation inducers. It is concluded that TPA increases the apoptotic action of As(2)O(3), an effect mediated by ERK activation and GSH depletion. However, the increase in apoptosis is only effective in non-differentiated cells.     

Chemical basis for inter-colonial aggression in the stingless bee Scaptotrigona bipunctata (Hymenoptera: Apidae)

Inter-colonial aggression was tested using three colonies of Scaptotrigona bipunctata in a natural setting when their nests were moved and by artificial contact between individuals. Examination of the cuticular lipids of individuals from two colonies kept under identical conditions showed clear differences in their cuticular hydrocarbon profiles. The cuticular lipids were a mixture of hydrocarbons (saturated and unsaturated alkanes and alkenes) within the range of C23-C29. The use of multivariate analysis (PCA and discriminant analysis) showed that seven of the identified surface compounds are enough to separate workers from colonies A and B from each other.     

Insect growth inhibition by tocotrienols and hydroquinones from Roldana barba-johannis

The methanol extract from the aerial parts of Roldana barba-johannis (Asteraceae) afforded sargachromenol, sargahydroquinoic acid, and sargaquinoic acid. These natural products and their corresponding acetylated and methylated derivatives showed insecticidal and insect growth regulatory activities against the Fall Armyworm [Spodoptera frugiperda J.E. Smith, (Lepidoptera: Noctuidae)], an important insect pest of corn. The most active compounds were sargachromenol and its acetylated derivative; sargahydroquinoic acid and its acetylated derivative; and a mixture of sargachromenol, sargahydroquinoic acid, and sargaquinoic acid (6:3:1) and the acetylated form of this mixture. All these compounds and mixtures had significant inhibitory effects between 5.0 and 20.0 ppm in diets. Most compounds were insecticidal to larvae, with lethal doses between 20 and 35 ppm. In addition, these substances also demonstrated scavenging properties toward 2,2-diphenyl-1-picrylhydrazyl radical in TLC autographic and spectrophotometric assays. These compounds appear to have selective effects on the pre-emergence metabolism of the insect. The results from these compounds were fully comparable in activity to those known natural insect growth inhibitors such as gedunin and methanol extracts of Cedrela salvadorensis and Yucca periculosa. These substances may be useful as natural insecticidal agents.     

Regulation of the human NBC3 Na+/HCO3- cotransporter by carbonic anhydrase II and PKA

Human NBC3 is an electroneutral Na⁺/HCO₃^– cotransporter expressed in heart, skeletal muscle, and kidney in which it plays an important role in HCO₃^– metabolism. Cytosolic enzyme carbonic anhydrase II (CAII) catalyzes the reaction CO₂ + H₂O HCO₃^– + H⁺ in many tissues. We investigated whether NBC3, like some Cl^–/HCO₃^– exchange proteins, could bind CAII and whether PKA could regulate NBC3 activity through modulation of CAII binding. CAII bound the COOH-terminal domain of NBC3 (NBC3Ct) with K_d = 101 nM; the interaction was stronger at acid pH. Cotransfection of HEK-293 cells with NBC3 and CAII recruited CAII to the plasma membrane. Mutagenesis of consensus CAII binding sites revealed that the D1135-D1136 region of NBC3 is essential for CAII/NBC3 interaction and for optimal function, because the NBC3 D1135N/D1136N retained only 29 ± 22% of wild-type activity. Coexpression of the functionally dominant-negative CAII mutant V143Y with NBC3 or addition of 100 µM 8-bromoadenosine to NBC3 transfected cells reduced intracellular pH (pH_i) recovery rate by 31 ± 3, or 38 ± 7%, respectively, relative to untreated NBC3 transfected cells. The effects were additive, together decreasing the pH_i recovery rate by 69 ± 12%, suggesting that PKA reduces transport activity by a mechanism independently of CAII. Measurements of PKA-dependent phosphorylation by mass spectroscopy and labeling with [-³²P]ATP showed that NBC3Ct was not a PKA substrate. These results demonstrate that NBC3 and CAII interact to maximize the HCO₃^– transport rate. Although PKA decreased NBC3 transport activity, it did so independently of the NBC3/CAII interaction and did not involve phosphorylation of NBC3Ct. pH regulation; bicarbonate transport; metabolon     

Etoposide stimulates 1,25-dihydroxyvitamin D3 differentiation activity,hormone binding and hormone receptor expression in HL-60 human promyelocytic cells

The simultaneous administration of the DNA topoisomerase II inhibitor etoposide (0.15 mM) and 1,25-dihydroxyvitamin D₃ (VD₃) (10 nM) synergistically induced the differentiation of HL-60 human promyelocytic leukemia cells. Similar results were obtained using U-937 human promonocytic cells, or the topoisomerase II inhibitors doxorubicin (15 nM) and mitoxantrone (2.5 nM). When sequential treatments were used, pre-incubation with VD₃ had little effect on the subsequent action of etoposide, while pre-incubation with etoposide greatly potentiated the subsequent action of VD₃. In addition, etoposide treatment stimulated VD₃ binding activity and increased VD₃ receptor mRNA and protein levels. The increase in hormone receptor expression may explain, at least in part, the capacity of topoisomerase inhibitors to potentiate the differentiation inducing activity of VD₃.     

A protein phosphatase 1 gamma (PP1γ) of the human protozoan parasite Trichomonas vaginalis is involved in proliferation and cell attachment to the host cell

In this work, evidence for a critical role of Trichomonas vaginalis protein phosphatase 1 gamma (TvPP1γ) in proliferation and attachment of the parasite to the mammalian cell is provided. Firstly, proliferation and attachment of T. vaginalis parasites to HeLa cells was blocked by calyculin A (CA), a potent PP1 inhibitor. Secondly, it was demonstrated that the enzyme activity of native and recombinant TvPP1γ proteins was inhibited by CA. Thirdly, reverse genetic studies confirmed that antisense oligonucleotides targeted to PP1γ but not PP1α or β inhibited proliferation and attachment of trichomonads CA-treated parasites underwent cytoskeletal modifications, including a lack of axostyle typical labelling, suggesting that cytoskeletal phosphorylation could be regulated by a CA-sensitive phosphatase where the role of PP1γ could not be ruled out. Analysis of subcellular distribution of TvPP1γ by cell fractionation and electron microscopy demonstrated the association between TvPP1γ and the cytoskeleton. The expression of adhesins, AP120 and AP65, at the cell surface was also inhibited by CA. The concomitant inhibition of expression of adhesins and changes in the cytoskeleton in CA-treated parasites suggest a specific role for PP1γ -dependent dephosphorylation in the early stages of the host-parasite interaction. Molecular modelling of TvPP1γ showed the conservation of residues critical for maintaining proper folding into the gross structure common to PP1 proteins. Taken together, these results suggest that TvPP1γ could be considered a potential novel drug target for treatment of trichomoniasis.     

Pollination: A key event controlling the expression of genes related to phytohormone biosynthesis during grapevine berry formation

Berry formation is the process of ovary conversion into a functional fruit, and is characterized by abrupt changes in the content of several phytohormones, associated with pollination and fertilization. Much effort has been made in order to improve our understanding of berry development, particularly from veraison to post-harvest time. However, the period of berry formation has been poorly investigated, despite its importance. Phytohormones are involved in the control of fruit formation; hence it is important to understand the regulation of their content at this stage. Grapevine is an excellent fleshy-fruit plant model since its fruits have particularities that differentiate them from those of commonly studied organisms. For instance, berries are prepared to cope with stress by producing several antioxidants and they are non-climacteric fruits. Also its genome is fully sequenced, which allows to identify genes involved in developmental processes. In grapevine, no link has been established between pollination and phytohormone biosynthesis, until recently. Here we highlight relevant findings regarding pollination effect on gene expression related to phytohormone biosynthesis, and present results showing how quickly this effect is achieved.