Inhibitory effect of genital photoreceptors on the activity of abdominal motoneurones in Papilio xuthus (Lepidoptera,Papilionidae)

The inhibitory effect of butterfly genital photoreceptors on the activities of abdominal motoneurones is described. In two (N1, N3) of the six lateral nerves (N1–6) belonging to the last abdominal ganglion, spontaneous motoneurone activity was inhibited by illumination of the genital photoreceptors. N1 and N3 innervate the ventral longitudinal muscles. N2 and N4, which supply branches to the spiracular muscles, were not inhibited. The results allow some of the properties of the circuits in the abdominal nervous system supplied by the genital photoreceptors to be inferred. Some possible functions of the photoreceptors are discussed.     

Effects of adenosine, 2-deoxyadenosine and N6-phenylisopropyladenosine on rat islet function and metabolism

Adenosine (1.0-100 mum). N(6)-phenylisopropyladenosine (0.1-10 mum) and 2-deoxyadenosine (10 mm) all produced a dose-dependent inhibition of glucose-stimulated insulin release. The inhibition of glucose-stimulated insulin release by adenosine and N(6)-phenylisopropyladenosine was abolished by 3-isobutyl-1-methylxanthine (0.1 mm), whereas 2-deoxyadenosine inhibited insulin release even in the presence of 3-isobutyl-1-methylxanthine. These adenosine nucleosides also inhibited the release of insulin induced by 4-methyl-2-oxopentanoate (20 mm), dl-glyceraldehyde (30 mm) and l-leucine (20 mm). Adenosine (10 mum). N(6)-phenylisopropyladenosine (10 mum) and 2-deoxyadenosine (10 mm) did not inhibit insulin biosynthesis or [U-(14)C]glucose oxidation at concentrations of the nucleosides that gave maximal inhibition of insulin release. However, adenosine, 2-deoxyadenosine and N(6)-phenylisopropyladenosine produced marked inhibition of the glucose-stimulated increases seen in islet cyclic AMP accumulation. Similar to its effects on insulin release, 3-isobutyl-1-methylxanthine (0.1 mm) antagonized the inhibitory effects of cyclic AMP accumulation produced by adenosine and N(6)-phenylisopropyladenosine, but had no effect on the inhibition of cyclic AMP accumulation seen with 2-deoxyadenosine. These results show that adenosine and its specifically modified analogues, 2-deoxyadenosine and N(6)-phenylisopropyladenosine, are strong inhibitors of insulin release from rat islets, a function that appears to be the consequence of their ability to inhibit the accumulation of cyclic AMP. It is proposed that the B cells, in common with many other tissues, may possess two different sites at which adenosine nucleosides interact to produce their biological effects; these are the so-called ;P' and ;R' sites first described by Londos & Wolff [(1977) Proc. Natl. Acad. Sci. U.S.A.74, 5482-5486].     

Ca2+, calmodulin, and protein dephosphorylation are required for GA-induced gene expression in petunia corolla

Gibberellic acid (GA₃) induces the expression of different genes, including chalcone synthase ( chs ) and gip , in detached petunia corollas. To initiate a study on gibberellin (GA)-signal transduction in this tissue, we examined the effect of agents that inhibit or promote specific steps in signal-transduction pathways. The calcium chelator 1,2- bis ( o -aminophenoxy)ethane N,N,N ' ,N '-tetraacetic acid (BAPTA) had no effect on GA-induced gene expression, while the calcium-channel blocker, ruthenium red (RR), inhibited the activation of the genes. The calmodulin antagonist N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (W-7) inhibited the induction of chs and gip by the hormone, and its analog, N-(6-aminohexyl)-1-naphthalenesulfonamide hydrochloride (W-5), had lower effect. The activation of chs and gip by GA₃ was completely blocked by the protein phosphatase inhibitor, okadaic acid (OA), and partially inhibited by the protein kinase inhibitor, 1-(5-isoquinoline-sulfonyl)- 2-methylpiperazine dihydrochloride (H-7). We suggest that Ca²⁺ from intracellular sources, calmodulin and protein dephosphorylation and phosphorylation are involved in GA-induced gene expression in petunia corollas.     

Neurotensin, bradykinin and somatostatin inhibit cAMP production in neuroblastoma N1E115 cells via both pertussis toxin sensitive and insensitive mechanisms

Neurotensin, bradykinin and somatostatin inhibited in a time- and concentration-dependent manner prostaglandin E1- or forskolin-stimulated cAMP production in neuroblastoma N1E115 cells. Cell treatment with 1 microgram/ml pertussis toxin for 6 hours reversed the inhibition elicited by peptides after short incubation periods (less than or equal to 1 min) but, in contrast, had no effect after longer incubation periods (greater than or equal to 3 min). Fluoroaluminate also inhibited prostaglandin E1-stimulated cAMP production in N1E115 cells, and this effect was not reversed by pertussis toxin. The 6 hour treatment with pertussis toxin was shown to be sufficient to ADP ribosylate virtually all of the 41 kD protein substrate corresponding to the alpha subunit of Gi. Protein kinase C activation with phorbol ester did not inhibit basal or stimulated cAMP production. Our data point to the existence of both pertussis toxin sensitive and insensitive mechanisms of neuropeptide-mediated inhibition of cAMP formation in N1E115 cells. The toxin insensitive response is not mediated by protein kinase C. The possibility is discussed that it results from the activation of a pertussis toxin insensitive G protein.     

H9N2 virus-derived M1 protein promotes H5N6 virus release in mammalian cells: Mechanism of avian influenza virus inter-species infection in humans

H5N6 highly pathogenic avian influenza virus (HPAIV) clade 2.3.4.4 not only exhibits unprecedented intercontinental spread in poultry, but can also cause serious infection in humans, posing a public health threat. Phylogenetic analyses show that 40% (8/20) of H5N6 viruses that infected humans carried H9N2 virus-derived internal genes. However, the precise contribution of H9N2 virus-derived internal genes to H5N6 virus infection in humans is unclear. Here, we report on the functional contribution of the H9N2 virus-derived matrix protein 1 (M1) to enhanced H5N6 virus replication capacity in mammalian cells. Unlike H5N1 virus-derived M1 protein, H9N2 virus-derived M1 protein showed high binding affinity for H5N6 hemagglutinin (HA) protein and increased viral progeny particle release in different mammalian cell lines. Human host factor, G protein subunit beta 1 (GNB1), exhibited strong binding to H9N2 virus-derived M1 protein to facilitate M1 transport to budding sites at the cell membrane. GNB1 knockdown inhibited the interaction between H9N2 virus-derived M1 and HA protein, and reduced influenza virus-like particles (VLPs) release. Our findings indicate that H9N2 virus-derived M1 protein promotes avian H5N6 influenza virus release from mammalian, in particular human cells, which could be a major viral factor for H5N6 virus cross-species infection.     

Structure and activity relationships of novel uracil derivatives as topical anti-inflammatory agents

In order to create novel, topical anti-inflammatory compounds exhibiting more potent activities than lead compound CX-659S (1), we designed and synthesized various derivatives of 1 focusing on the uracil N(1)- and N(3)-substituents, and evaluated their anti-inflammatory activities via inhibition of the picryl chloride-induced contact hypersensitivity reaction (CHR) in mice. In the course of our structure and activity relationship study, we found that compounds 6k, 6q, and 6r inhibited by approximately 50% the CHR, at 0.1 mg/ear. These activities were essentially equipotent with that of Tacrolimus, a strong immunosuppressant.     

Role of the conserved NPxxY motif of the 5-HT2A receptor in determining selective interaction with isoforms of ADP-ribosylation factor (ARF)

In this study we have shown that N376 to D mutation in the conserved NPxxY motif within the carboxy terminal tail domain (CT) of the 5-HT2A receptor alters the binding preference of GST-fusion protein constructs of the CT domain from ARF1 to an alternative isoform, ARF6. These findings were corroborated by experiments investigating co-immunoprecipitation of the wild type (WT) and N376D mutant of the 5-HT2A receptor with ARF1 or 6 or dominant negative ARF1/6 constructs co-expressed in COS7 cells. In functional assays of 5-HT-induced phospholipase D (PLD) activation responses of the WT receptor were inhibited by a dominant negative mutant of ARF1 but not ARF6, whereas responses of the N376D mutant were strongly inhibited by negative mutant ARF6. No equivalent effect of the ARF mutants was seen on phospholipase C activation. In experiments assaying 5-HT-induced increases in [35S]GTPgammaS binding to ARF 1/6 immunoprecipitates as a measure of ARF activation, increased ARF6 activation was seen only with the mutant receptor. When cellular PLD responses of other NPxxY- or a DPxxY-containing GPCRs were measured in the presence of dominant negative ARF1/6 constructs, the majority, but not all, fitted the pattern exemplified by the 5-HT2A receptor and its N376D mutant. These data suggest that the presence of the N or a D in this highly conserved motif is an important, but not exclusive, determinant of which ARF isoform interacts with the GPCR.     

Modulator of intracellular Ca(2+), thapsigargin, interferes with in vitro secretion of cytokines and nitric oxide

Interference of thapsigargin (TG), an inhibitor of endoplasmic reticulum Ca(2+) ATPase, with immune reactivity of murine macrophages was investigated under conditions in vitro. The activation of cells with lipopolysaccharide (LPS), interferon-(gamma) (IFN-(gamma)), and with acyclic nucleoside phosphonate N(6)-isobutyl-9-[2-(phosphonomethoxy)ethyl]- 2,6-diaminopurine (N(6)-isobutyl-PMEDAP) resulted in enhanced production of cytokines TNF-alpha, IL-10, chemokines RANTES/CCL5 and MIP-1alpha/CCL3, as well as in substantially augmented production of nitric oxide (NO) triggered by IFN-(gamma). The effects were in a dual mode of action influenced by TG (1 microM). While TG upregulated secretion of TNF-alpha, it inhibited secretion of IL-10 and RANTES. The immune-stimulated secretion of MIP-1alpha remained virtually unaffected, though TG on its own activated expression of MIP-1alpha in macrophages. The high-output NO production induced by IFN-(gamma), high concentrations of LPS, or by combination of IFN-(gamma) plus LPS or N(6)-isobutyl-PMEDAP was inhibited by TG. On the other hand, production of NO which was marginally activated by low concentration of LPS was upregulated by TG.     

Matrix NADH dehydrogenases of plant mitochondria and sites of quinone reduction by complex I.

In order to distinguish the pathways involved in the oxidation of matrix NADH in plant mitochondria, the oxidation of NADH and nicotinamide hypoxanthine dinucleotide (reduced form) was investigated in submitochondrial particles prepared from beetroot (Beta vulgaris L. cv. Derwent Globe) and soybeans (Glycine max L. cv. Bragg). Nicotinamide-hypoxanthine-dinucleotide(reduced form)-oxidase activity was more strongly inhibited by rotenone than the NADH-oxidase activity but both of the rotenone-inhibited activities could be stimulated by adding ubiquinone-1. The corresponding ubiquinone-1-reductase activities were inhibited by rotenone (to 69%) and further inhibited by N,N'-dicyclohexylcarbodiimide (to 79%), whilst the K3Fe(CN)6-reductase activities were not sensitive to either rotenone or N,N'-dicyclohexylcarbodiimide. Immunological analysis of mitochondrial proteins using an antiserum raised against purified beetroot complex I indicated very few differences between soybean and fresh and aged beetroot mitochondria, despite their varying sensitivities to rotenone. We confirm that there are two dehydrogenases capable of oxidising internal NADH and that only one of these, namely complex I, is inhibited by rotenone. Further, we conclude that complex I has two potential sites of quinone reduction, both sensitive to N,N'-dicyclohexycarbodiimide inhibition but only one of which is sensitive to rotenone inhibition.     

Mechanism-based irreversible inhibitors of cytokinin dehydrogenase

The effects of three N(6)-substituted aminopurine derivatives containing either allenic or acetylenic side-chains on in vitro and in vivo cytokinin dehydrogenase (CKX; EC 1.5.99.12) activities were determined. At concentrations < or = 100 microM, the acetylenic derivative (HA-2) had no effect on in vitro CKX activity. In contrast, the two allenic derivatives (HA-1, HA-8) inhibited in vitro CKX activity in a dose-dependent manner with 50% inhibition occurring at HA-1 and HA-8 concentrations of 9.0 and 0.4 microM (respectively). HA-8 inhibited the degradation of both the free bases and ribosides of N6-(2-isopentenyl)adenine and zeatin. Pretreatment with HA-8 inhibited CKX activity in both a time- and concentration-dependent manner. In contrast to the reversible phenylurea inhibitor N-(chloro-4-pyridyl)-N'-phenylurea, inhibition of CKX activity by HA-8 was not relieved by 24 h of dialysis. Both HA-1 and HA-8 (but not HA-2) inhibited the metabolism of exogenous [3H]-N(6)-(2-isopentenyl)adenosine in excised aseptic potato (Solanum tuberosum) leaves. These results demonstrate that HA-8 is a mechanism-based irreversible (suicide) inhibitor of CKX and indicate that it may be useful in determining the role of CKX in cytokinin homeostasis in planta.     

Glycyrrhizin exerts antioxidative effects in H5N1 influenza A virus-infected cells and inhibits virus replication and pro-inflammatory gene expression

Glycyrrhizin is known to exert antiviral and anti-inflammatory effects. Here, the effects of an approved parenteral glycyrrhizin preparation (Stronger Neo-Minophafen C) were investigated on highly pathogenic influenza A H5N1 virus replication, H5N1-induced apoptosis, and H5N1-induced pro-inflammatory responses in lung epithelial (A549) cells. Therapeutic glycyrrhizin concentrations substantially inhibited H5N1-induced expression of the pro-inflammatory molecules CXCL10, interleukin 6, CCL2, and CCL5 (effective glycyrrhizin concentrations 25 to 50 μg/ml) but interfered with H5N1 replication and H5N1-induced apoptosis to a lesser extent (effective glycyrrhizin concentrations 100 μg/ml or higher). Glycyrrhizin also diminished monocyte migration towards supernatants of H5N1-infected A549 cells. The mechanism by which glycyrrhizin interferes with H5N1 replication and H5N1-induced pro-inflammatory gene expression includes inhibition of H5N1-induced formation of reactive oxygen species and (in turn) reduced activation of NFκB, JNK, and p38, redox-sensitive signalling events known to be relevant for influenza A virus replication. Therefore, glycyrrhizin may complement the arsenal of potential drugs for the treatment of H5N1 disease.     

Severe acute respiratory syndrome coronavirus open reading frame (ORF) 3b, ORF 6, and nucleocapsid proteins function as interferon antagonists

The severe acute respiratory syndrome coronavirus (SARS-CoV) is highly pathogenic in humans, with a death rate near 10%. This high pathogenicity suggests that SARS-CoV has developed mechanisms to overcome the host innate immune response. It has now been determined that SARS-CoV open reading frame (ORF) 3b, ORF 6, and N proteins antagonize interferon, a key component of the innate immune response. All three proteins inhibit the expression of beta interferon (IFN-beta), and further examination revealed that these SARS-CoV proteins inhibit a key protein necessary for the expression of IFN-beta, IRF-3. N protein dramatically inhibited expression from an NF-kappaB-responsive promoter. All three proteins were able to inhibit expression from an interferon-stimulated response element (ISRE) promoter after infection with Sendai virus, while only ORF 3b and ORF 6 proteins were able to inhibit expression from the ISRE promoter after treatment with interferon. This indicates that N protein inhibits only the synthesis of interferon, while ORF 3b and ORF 6 proteins inhibit both interferon synthesis and signaling. ORF 6 protein, but not ORF 3b or N protein, inhibited nuclear translocation but not phosphorylation of STAT1. Thus, it appears that these three interferon antagonists of SARS-CoV inhibit the interferon response by different mechanisms.     

Human Immunodeficiency Virus Type 1 (HIV-1) Infection and Transcytosis Activity of a HIV-1 Susceptible Clone from HeLa Cell

In order to clarify the transmission process of human immunodeficiency virus type 1 (HIV-1) through the epithelial cell barrier, HeLa cells susceptible and non-susceptible to HIV-1 were cloned and designated as P6 HeLa and N7 HeLa cells, respectively. P6 HeLa cells could be infected with the LAI strain of HIV-1 and mediated HIV-1 transcytosis. In contrast, N7 HeLa cells exhibited neither HIV-1 infection nor transcytosis. CD4 and galactosylceramide as the receptors for HIV-1 were not detected on P6 HeLa cells, although an anti-CD4 monoclonal antibody (mAb) blocked HIV-1 infection. Since HIV-1-infected P6 HeLa cells exhibited no fusion and survived, we speculated that the P6 HeLa cells expressed molecules other than CD4 which facilitated HIV-1 infection. Two mAbs (A-14 ITK and C57 a9-9) which inhibited the HIV-1 infection of P6 HeLa cells were generated. Each mAb recognized distinct molecule(s) as shown by Western blotting. Transcytosis by the P6 HeLa cells was inhibited by C57 a9-9 but not by A-14 ITK or anti-CD4 mAb. Both infection and transcytosis may be responsible for HIV-1 transmission through epithelial cells in a complex manner. Although infection and transcytosis occurred via different mechanisms, the molecule(s) recognized by C57 a9-9 mAb may be associated with both processes.     

ARF6 is required for growth factor- and rac-mediated membrane ruffling in macrophages at a stage distal to rac membrane targeting

Activation of Rac1, a member of the Rho family of GTPases, is associated with multiple cellular responses, including membrane ruffling and focal complex formation. The mechanisms by which Rac1 is coupled to these functional responses are not well understood. It was recently shown that ARF6, a GTPase implicated in cytoskeletal alterations and a membrane recycling pathway, is required for Rac1-dependent phagocytosis in macrophages (Q. Zhang et al., J. Biol. Chem. 273:19977-19981, 1998). To determine whether ARF6 is required for Rac1-dependent cytoskeletal responses in macrophages, we expressed wild-type (WT) or guanine nucleotide binding-deficient alleles (T27N) of ARF6 in macrophages coexpressing activated alleles of Rac1 (Q61L) or Cdc42 (Q61L) or stimulated with colony-stimulating factor 1 (CSF-1). Expression of ARF6 T27N but not ARF6 WT inhibited ruffles mediated by Rac1 Q61L or CSF-1. In contrast, expression of ARF6 T27N did not inhibit Rac1 Q61L-mediated focal complex formation and did not impair Cdc42 Q61L-mediated filopodial formation. Cryoimmunogold electron microscopy demonstrated the presence of ARF6 in membrane ruffles induced by either CSF-1 or Rac1 Q61L. Addition of CSF-1 to macrophages led to the redistribution of ARF6 from the interior of the cell to the plasma membrane, suggesting that this growth factor triggers ARF6 activation. Direct targeting of Rac1 to the plasma membrane did not bypass the blockade in ruffling induced by ARF6 T27N, indicating that ARF6 regulates a pathway leading to membrane ruffling that occurs after the activation and membrane association of Rac. These data demonstrate that intact ARF6 function is required for coupling activated Rac to one of several effector pathways and suggest that a principal function of ARF6 is to coordinate Rac activation with plasma membrane-based protrusive events.     

Rho family GTPases and neuronal growth cone remodelling: relationship between increased complexity induced by Cdc42Hs, Rac1, and acetylcholine and collapse induced by RhoA and lysophosphatidic acid.

Rho family GTPases have been assigned important roles in the formation of actin-based morphologies in nonneuronal cells. Here we show that microinjection of Cdc42Hs and Rac1 promoted formation of filopodia and lamellipodia in N1E-115 neuroblastoma growth cones and along neurites. These actin-containing structures were also induced by injection of Clostridium botulinum C3 exoenzyme, which abolishes RhoA-mediated functions such as neurite retraction. The C3 response was inhibited by coinjection with the dominant negative mutant Cdc42Hs(T17N), while the Cdc42Hs response could be competed by coinjection with RhoA. We also demonstrate that the neurotransmitter acetylcholine (ACh) can induce filopodia and lamellipodia on neuroblastoma growth cones via muscarinic ACh receptor activation, but only when applied in a concentration gradient. ACh-induced formation of filopodia and lamellipodia was inhibited by preinjection with the dominant negative mutants Cdc42Hs(T17N) and Rac1(T17N), respectively. Lysophosphatidic acid (LPA)-induced neurite retraction, which is mediated by RhoA, was inhibited by ACh, while C3 exoenzyme-mediated neurite outgrowth was inhibited by injection with Cdc42Hs(T17N) or Rac1(T17N). Together these results suggest that there is competition between the ACh- and LPA-induced morphological pathways mediated by Cdc42Hs and/or Rac1 and by RhoA, leading to either neurite development or collapse.     

Glucose transport in Achlya: characterization and possible regulatory aspects

The freshwater fungus Achlya transported D-(+)glucose (glucose) and 2-deoxy-D-glucose (deoxyglucose) by an energy-related system. Their transport4 was inhibited by uncouplers of metabolic energy such as 2,4-dinitrophenol, cyanide, azide, and carbonylcyanide-p-chlorophenylhydrazone. Besides inhibiting each other, glucose and deoxyglucose transport was inhibited by D-(+)galactose, D-(+)mannose, and D-(+)xylose. Many other sugars tested failed to inhibit glucose transport implying a certain degree of specificity. Glucose transport was pH (optimum at 6.5) and temperature (optimum at 30-40 degrees C) dependent. Glucose transport was also inhibited by citrate, N6-substituted adenines (cytokinins), and iodine. None of these agents penetrated the cell membrane within the brief (1-3-min) period in which glucose transport was measured. In every case, transport was inhibited within 10 s (the shortest time in which measurements could be made). When cells were osmotically shocked to release a cell-wall membrane phosphorylated proteoglycan (PPG), they became incapable of transporting glucose for several hours until new PPG material was reisolable from the membrane by osmotic-shock treatment. The osmotically shocked cells could not transport glucose or deoxyglucose. No glucose-binding protein was detected in the shock fluid. Practically all of the glucose transported within 1-2 min was recovered as glucose-6-phosphate. No other phosphorylated sugar was detected suggesting that glucose may be phosphorylated in transport. Related studies have shown that citrate removed calcium bound by PPG; N6-substituted adenines were bound by PPG while three polyphosphorylated dinucleosides, HS3, HS2, and HS1, were displaced from it. Iodine formed stable complexes with the HS compounds. All of these agents inhibited glucose transport without entering the cell. It is therefore possible that HS compounds, calcium and PPG may be involved in maintaining the cell membrane in proper form for glucose transport.     

Inhibition of ALDH3A1-catalyzed oxidation by chlorpropamide analogues

In our efforts to identify agents that would specifically inhibit ALDH3A1, we had previously studied extensively the effect of an N(1)-alkyl, an N(1)-methoxy, and several N(1)-hydroxy-substituted ester derivatives of chlorpropamide on the catalytic activities of ALDH3A1s derived from human normal stomach mucosa (nALDH3A1) and human tumor cells (tALDH3A1), and of two recombinant aldehyde dehydrogenases, viz. human rALDH1A1 and rALDH2. The N(1)-methoxy analogue of chlorpropamide, viz. 4-chloro-N-methoxy-N-[(propylamino)carbonyl]benzenesulfonamide (API-2), was found to be a relatively selective and potent inhibitor of tALDH3A1-catalyzed oxidation as compared to its ability to inhibit nALDH3A-catalyzed oxidation, but even more potently inhibited ALDH2-catalyzed oxidation, whereas an ester analogue, viz. (acetyloxy)[(4-chlorophenyl)sulfonyl]carbamic acid 1,1-dimethylethyl ester (NPI-2), selectively inhibited tALDH3A1-catalyzed oxidation as compared to its ability to inhibit nALDH3A1-, ALDH1A1- and ALDH2-catalyzed oxidations, and this inhibition was apparently irreversible. Three additional chlorpropamide analogues, viz. 4-chloro-N,O-bis(ethoxycarbonyl)-N-hydroxybenzenesulfonamide (NPI-4), N,O-bis(carbomethoxy)methanesulfohydroxamic acid (NPI-5), and 2-[(ethoxycarbonyl)oxy]-1,2-benzisothiazol-3(2H)-one 1,1-dioxide (NPI-6), were evaluated in the present investigation. Quantified were NAD-linked oxidation of benzaldehyde catalyzed by nALDH3A1 and tALDH3A1, and NAD-linked oxidation of acetaldehyde catalyzed by rALDH1A1 and rALDH2, all at 37 degrees C and pH 8.1, and in the presence and absence of inhibitor. NPI-4, NPI-5 and NPI-6 were not substrates for the oxidative reactions catalyzed by any of the ALDHs studied. Oxidative reactions catalyzed by the ALDH3A1s, rALDH1A1 and rALDH2 were each inhibited by NPI-4 and NPI-5. NPI-6 was a poor inhibitor of nALDH3A1- and tALDH3A1-catalyzed oxidations, but was a relatively potent inhibitor of rALDH1A1- and rALDH2-catalyzed oxidations. In all cases, inhibition of ALDH-catalyzed oxidation was directly related to the product of inhibitor concentration and preincubation (enzyme+inhibitor) time. As judged by the product values (microMxmin) required to effect 50% inhibition (IC(50)): (1) nALDH3A1 and tALDH3A1 were essentially equisensitive to inhibition by NPI-4 and NPI-5, and both enzymes were poorly inhibited by NPI-6; (2) rALDH1A1 was, relative to the ALDH3A1s, slightly more sensitive to inhibition by NPI-4 and NPI-5, and far more sensitive to inhibition by NPI-6; and (3) rALDH1A1 was, relative to rALDH2, essentially equisensitive to inhibition by NPI-5, whereas, it was slightly more sensitive to inhibition by NPI-4 and NPI-6.     

The ability of salts to inhibit the reaction between periodate anions and ovotransferrin.

N1-Acetylspermidine is not detectable in rat heart, but its content greatly increases after a single injection of isoprenaline (10 mg/kg), reaching a concentration of about 10 nmol/g of tissue 4 h after the treatment. Part of the accumulated N1-acetylspermidine was split to putrescine. Isoprenaline also caused an increase of N1-acetylspermidine in the spleen, where its concentration increased 3.5-fold 6 h after the catecholamine. The accumulation of N1-acetylspermidine was dependent on the dose of isoprenaline in both the heart and the spleen, and was strongly inhibited by beta-antagonists and inhibitors of protein synthesis.     

Potyviral 6K2 protein long-distance movement and symptom-induction functions are independent and host-specific

Deletion of various portions, or insertion of six histidine residues (6xHis) into various positions of the membrane-bound 6K2 protein (53 amino acids) of Potato virus A (PVA, genus Potyvirus), inhibited systemic infection in Nicotiana tabacum and N. benthamiana plants. However, a spontaneous mutation (Gly2Cys) that occurred in 6K2 adjacent to the 6xHis insert placed between Ser1 and Gly2 enabled systemic infection in a single N. benthamiana plant. No symptoms were observed, but virus titers were similar to the symptom-inducing wild-type (wt) PVA. N. tabacum plants were not systemically infected, albeit virus propagation was observed in inoculated protoplasts. The 6xHis/Gly2Cys mutant was reconstructed in vitro and serially propagated by mechanical inoculation in N. benthamiana. Following the third passage, a novel viral mutant appeared, lacking the last four His residues of the insert, as well as the Gly2 and Thr3 of 6K2. It infected N. tabacum plants systemically, and in the systemically infected N. benthamiana leaves, vein chlorosis and mild yellowing symptoms were observed, typical of wt PVA infection. The mutant virus accumulated to titers similar to wt PVA in both hosts. These results show that the PVA 6K2 protein affects viral long-distance movement and symptom induction independently and in a host-specific manner.