O PAPEL DA COMUNICA����O FACE A FACE NAS ORGANIZA����ES NO CONTEXTO DA SOCIEDADE MIDIATIZADA

Este estudo trata da comunica����o face a face nas organiza����es sob diferentes abordagens te��ricas. Considera a perspectiva da simultaneidade dos meios, j�� que as empresas utilizam diversos canais para dialogar com seus p��blicos de interesse. Leva em conta o fen��meno da midiatiza����o, que reestrutura o modo como as pessoas se relacionam na sociedade contempor��nea. O objetivo geral da pesquisa �� sistematizar papeis potencialmente exercidos pela intera����o face a face e conhecer algumas circunst��ncias que envolvem sua pr��tica nas organiza����es. Por se tratar de uma tese te��rica, a pesquisa bibliogr��fica se apresenta como um dos principais procedimentos metodol��gicos; an��lises de casos emp��ricos e um estudo de caso desenvolvido na Embrapa Pantanal constituem situa����es ilustrativas. Conclui-se que a comunica����o face a face nas empresas ocorre de forma simult��nea e combinada a outros canais de comunica����o, por��m, ela proporciona resultados pr��ticos e filos��ficos ainda pouco explorados. �� rara a utiliza����o estrat��gica de contatos presenciais como mecanismo para estabelecer relacionamentos, conhecer as rea����es alheias e ajustar a comunica����o, aliar o discurso corporativo ��s pr��ticas empresariais e avaliar o contexto onde se desenvolvem as intera����es, o que pode ser decisivo para a comunica����o organizacional.

O DISCURSO AMBIENTAL JORNAL��STICO, POL��TICO E EMPRESARIAL EM ��MBITO LOCAL: uma proposta de an��lise

Esta tese tem como tema principal o discurso ambiental local. Trata-se de uma proposta de an��lise, que pode ser aplicada a este tipo de discurso em qualquer localidade, e um estudo de caso, que aplicou a an��lise proposta na tese. O objetivo geral desta pesquisa foi analisar a informa����o local sobre meio ambiente veiculada por meio dos discursos jornal��stico, pol��tico e empresarial, estabelecendo rela����es hist��ricas, sociais e ideol��gicas do discurso ambiental que permeiam as tr��s ��reas em quest��o. A metodologia incluiu tr��s fases: pesquisa bibliogr��fica e revis��o de literatura sobre os principais conceitos levantados no trabalho; levantamento de discursos locais dispon��veis na m��dia para an��lise no estudo de caso; an��lise dos discursos a partir de protocolo elaborado com base na An��lise do Discurso de linha francesa. O estudo de caso traz a an��lise do discurso ambiental no munic��pio de Frutal-MG, onde hoje est��o em andamento v��rias pesquisas e projetos na ��rea ambiental. A principal conclus��o do trabalho confirma a hip��tese da pesquisa de que a informa����o ambiental fruto dos discursos pol��tico, jornal��stico e empresarial em ��mbito local tem car��ter predominantemente situacional, mercadol��gico e propagand��stico, pouco focada em conscientizar e educar e com ��nfase em interesses comerciais e eleitorais e na resolu����o de problemas emergenciais.

Broccoli sprouts: An exceptionally rich source of inducers of enzymes that protect against���chemical���carcinogens

Induction of phase 2 detoxication enzymes [e.g., glutathione transferases, epoxide hydrolase, NAD(P)H: quinone reductase, and glucuronosyltransferases] is a powerful strategy for achieving protection against carcinogenesis, mutagenesis, and other forms of toxicity of electrophiles and reactive forms of oxygen. Since consumption of large quantities of fruit and vegetables is associated with a striking reduction in the risk of developing a variety of malignancies, it is of interest that a number of edible plants contain substantial quantities of compounds that regulate mammalian enzymes of xenobiotic metabolism. Thus, edible plants belonging to the family Cruciferae and genus Brassica (e.g., broccoli and cauliflower) contain substantial quantities of isothiocyanates (mostly in the form of their glucosinolate precursors) some of which (e.g., sulforaphane or 4-methylsulfinylbutyl isothiocyanate) are very potent inducers of phase 2 enzymes. Unexpectedly, 3-day-old sprouts of cultivars of certain crucifers including broccoli and cauliflower contain 10���100 times higher levels of glucoraphanin (the glucosinolate of sulforaphane) than do the corresponding mature plants. Glucosinolates and isothiocyanates can be efficiently extracted from plants, without hydrolysis of glucosinolates by myrosinase, by homogenization in a mixture of equal volumes of dimethyl sulfoxide, dimethylformamide, and acetonitrile at ���50��C. Extracts of 3-day-old broccoli sprouts (containing either glucoraphanin or sulforaphane as the principal enzyme inducer) were highly effective in reducing the incidence, multiplicity, and rate of development of mammary tumors in dimethylbenz(a)anthracene-treated rats. Notably, sprouts of many broccoli cultivars contain negligible quantities of indole glucosinolates, which predominate in the mature vegetable and may give rise to degradation products (e.g., indole-3-carbinol) that can enhance tumorigenesis. Hence, small quantities of crucifer sprouts may protect against the risk of cancer as effectively as much larger quantities of mature vegetables of the same variety.

CP12 provides a new mode of light regulation of Calvin cycle activity in higher���plants

CP12 is a small nuclear encoded chloroplast protein of higher plants, which was recently shown to interact with NAD(P)H���glyceraldehyde-3-phosphate dehydrogenase (GAPDH; EC 1.2.1.13), one of the key enzymes of the reductive pentosephosphate cycle (Calvin cycle). Screening of a pea cDNA library in the yeast two-hybrid system for proteins that interact with CP12, led to the identification of a second member of the Calvin cycle, phosphoribulokinase (PRK; EC 2.7.1.19), as a further specific binding partner for CP12. The exchange of cysteines for serines in CP12 demonstrate that interaction with PRK occurs at the N-terminal peptide loop of CP12. Size exclusion chromatography and immunoprecipitation assays reveal the existence of a stable 600-kDa PRK/CP12/GAPDH complex in the stroma of higher plant chloroplasts. Its stoichiometry is proposed to be of two N-terminally dimerized CP12 molecules, each carrying one PRK dimer on its N terminus and one A2B2 complex of GAPDH subunits on the C-terminal peptide loop. Incubation of the complex with NADP or NADPH, in contrast to NAD or NADH, causes its dissociation. Assays with the stromal 600-kDa fractions in the presence of the four different nicotinamide-adenine dinucleotides indicate that PRK activity depends on complex dissociation and might be further regulated by the accessible ratio of NADP/NADPH. From these results, we conclude that light regulation of the Calvin cycle in higher plants is not only via reductive activation of different proteins by the well-established ferredoxin/thioredoxin system, but in addition, by reversible dissociation of the PRK/CP12/GAPDH complex, mediated by NADP(H).

Requirement of poly(ADP-ribose) polymerase in recovery from DNA damage in mice and in���cells

Poly(ADP-ribose) polymerase [PARP; NAD+ ADP-ribosyltransferase; NAD+: poly(adenosine-diphosphate-d-ribosyl)-acceptor ADP-d-ribosyltransferase, EC 2.4.2.30] is a zinc-finger DNA-binding protein that detects specifically DNA strand breaks generated by genotoxic agents. To determine its biological function, we have inactivated both alleles by gene targeting in mice. Treatment of PARP���/��� mice either by the alkylating agent N-methyl-N-nitrosourea (MNU) or by ��-irradiation revealed an extreme sensitivity and a high genomic instability to both agents. Following whole body ��-irradiation (8 Gy) mutant mice died rapidly from acute radiation toxicity to the small intestine. Mice-derived PARP���/��� cells displayed a high sensitivity to MNU exposure: a G2/M arrest in mouse embryonic fibroblasts and a rapid apoptotic response and a p53 accumulation were observed in splenocytes. Altogether these results demonstrate that PARP is a survival factor playing an essential and positive role during DNA damage recovery.

Crystal structure of SQD1, an enzyme involved in the biosynthesis of the plant sulfolipid headgroup donor UDP-sulfoquinovose

The SQD1 enzyme is believed to be involved in the biosynthesis of the sulfoquinovosyl headgroup of plant sulfolipids, catalyzing the transfer of SO3��� to UDP-glucose. We have determined the structure of the complex of SQD1 from Arabidopsis thaliana with NAD+ and the putative substrate UDP-glucose at 1.6-��� resolution. Both bound ligands are completely buried within the binding cleft, along with an internal solvent cavity which is the likely binding site for the, as yet, unidentified sulfur-donor substrate. SQD1 is a member of the short-chain dehydrogenase/reductase (SDR) family of enzymes, and its structure shows a conservation of the SDR catalytic residues. Among several highly conserved catalytic residues, Thr-145 forms unusually short hydrogen bonds with both susceptible hydroxyls of UDP-glucose. A His side chain may also be catalytically important in the sulfonation.

A link between protein structure and enzyme catalyzed hydrogen���tunneling

We present evidence that the size of an active site side chain may modulate the degree of hydrogen tunneling in an enzyme-catalyzed reaction. Primary and secondary kH/kT and kD/kT kinetic isotope effects have been measured for the oxidation of benzyl alcohol catalyzed by horse liver alcohol dehydrogenase at 25��C. As reported in earlier studies, the relationship between secondary kH/kT and kD/kT isotope effects provides a sensitive probe for deviations from classical behavior. In the present work, catalytic efficiency and the extent of hydrogen tunneling have been correlated for the alcohol dehydrogenase-catalyzed hydride transfer among a group of site-directed mutants at position 203. Val-203 interacts with the opposite face of the cofactor NAD+ from the alcohol substrate. The reduction in size of this residue is correlated with diminished tunneling and a two orders of magnitude decrease in catalytic efficiency. Comparison of the x-ray crystal structures of a ternary complex of a high-tunneling (Phe-93 ��� Trp) and a low-tunneling (Val-203 ��� Ala) mutant provides a structural basis for the observed effects, demonstrating an increase in the hydrogen transfer distance for the low-tunneling mutant. The Val-203 ��� Ala ternary complex crystal structure also shows a hyperclosed interdomain geometry relative to the wild-type and the Phe-93 ��� Trp mutant ternary complex structures. This demonstrates a flexibility in interdomain movement that could potentially narrow the distance between the donor and acceptor carbons in the native enzyme and may enhance the role of tunneling in the hydride transfer reaction.

The synthesis of ATP by glycolytic enzymes in the postsynaptic density and the effect of endogenously generated nitric���oxide

The major contribution of this paper is the finding of a glycolytic source of ATP in the isolated postsynaptic density (PSD). The enzymes involved in the generation of ATP are glyceraldehyde-3-phosphate dehydrogenase (G3PD) and phosphoglycerate kinase (PGK). Lactate dehydrogenase (LDH) is available for the regeneration of NAD+, as well as aldolase for the regeneration of glyceraldehyde-3-phosphate (G3P). The ATP was shown to be used by the PSD Ca2+/calmodulin-dependent protein kinase and can probably be used by two other PSD kinases, protein kinase A and protein kinase C. We confirmed by immunocytochemistry the presence of G3PD in the PSD and its binding to actin. Also present in the PSD is NO synthase, the source of NO. NO increases the binding of NAD, a G3PD cofactor, to G3PD and inhibits its activity as also found by others. The increased NAD binding resulted in an increase in G3PD binding to actin. We confirmed the autophosphorylation of G3PD by ATP, and further found that this procedure also increased the binding of G3PD to actin. ATP and NO are connected in that the formation of NO from NOS at the PSD resulted, in the presence of NAD, in a decrease of ATP formation in the PSD. In the discussion, we raise the possible roles of G3PD and of ATP in protein synthesis at the PSD, the regulation by NO, as well as the overall regulatory role of the PSD complex in synaptic transmission.

A functional homolog of a yeast tRNA splicing enzyme is conserved in higher eukaryotes and in Escherichia coli

tRNA splicing in the yeast Saccharomyces cerevisiae requires an endonuclease to excise the intron, tRNA ligase to join the tRNA half-molecules, and 2���-phosphotransferase to transfer the splice junction 2���-phosphate from ligated tRNA to NAD, producing ADP ribose 1���������2������ cyclic phosphate (Appr>p). We show here that functional 2���-phosphotransferases are found throughout eukaryotes, occurring in two widely divergent yeasts (Candida albicans and Schizosaccharomyces pombe), a plant (Arabidopsis thaliana), and mammals (Mus musculus); this finding is consistent with a role for the enzyme, acting in concert with ligase, to splice tRNA or other RNA molecules. Surprisingly, functional 2���-phosphotransferase is found also in the bacterium Escherichia coli, which does not have any known introns of this class, and does not appear to have a ligase that generates junctions with a 2���-phosphate. Analysis of the database shows that likely members of the 2���-phosphotransferase family are found also in one other bacterium (Pseudomonas aeruginosa) and two archaeal species (Archaeoglobus fulgidus and Pyrococcus horikoshii). Phylogenetic analysis reveals no evidence for recent horizontal transfer of the 2���-phosphotransferase into Eubacteria, suggesting that the 2���-phosphotransferase has been present there since close to the time that the three kingdoms diverged. Although 2���-phosphotransferase is not present in all Eubacteria, and a gene disruption experiment demonstrates that the protein is not essential in E. coli, the continued presence of 2���-phosphotransferase in Eubacteria over large evolutionary times argues for an important role for the protein.

Characterization of the Saccharomyces cerevisiae ERG26 gene encoding the C-3 sterol dehydrogenase (C-4 decarboxylase) involved in sterol biosynthesis

All but two genes involved in the ergosterol biosynthetic pathway in Saccharomyces cerevisiae have been cloned, and their corresponding mutants have been described. The remaining genes encode the C-3 sterol dehydrogenase (C-4 decarboxylase) and the 3-keto sterol reductase and in concert with the C-4 sterol methyloxidase (ERG25) catalyze the sequential removal of the two methyl groups at the sterol C-4 position. The protein sequence of the Nocardia sp NAD(P)-dependent cholesterol dehydrogenase responsible for the conversion of cholesterol to its 3-keto derivative shows 30% similarity to a 329-aa Saccharomyces ORF, YGL001c, suggesting a possible role of YGL001c in sterol decarboxylation. The disruption of the YGL001c ORF was made in a diploid strain, and the segregants were plated onto sterol supplemented media under anaerobic growth conditions. Segregants containing the YGL001c disruption were not viable after transfer to fresh, sterol-supplemented media. However, one segregant was able to grow, and genetic analysis indicated that it contained a hem3 mutation. The YGL001c (ERG26) disruption also was viable in a hem 1�� strain grown in the presence of ergosterol. Introduction of the erg26 mutation into an erg1 (squalene epoxidase) strain also was viable in ergosterol-supplemented media. We demonstrated that erg26 mutants grown on various sterol and heme-supplemented media accumulate nonesterified carboxylic acid sterols such as 4��,14��-dimethyl-4��-carboxy-cholesta-8,24-dien-3��-ol and 4��-methyl-4��-carboxy-cholesta-8,24-dien-3��-ol, the predicted substrates for the C-3 sterol dehydrogenase. Accumulation of these sterol molecules in a heme-competent erg26 strain results in an accumulation of toxic-oxygenated sterol intermediates that prevent growth, even in the presence of exogenously added sterol.

Inosine-5���-Monophosphate Dehydrogenase Is a Rate-determining Factor for p53-dependent Growth Regulation

We have proposed that reduced activity of inosine-5���-monophosphate dehydrogenase (IMPD; IMP:NAD oxidoreductase, EC 1.2.1.14), the rate-limiting enzyme for guanine nucleotide biosynthesis, in response to wild-type p53 expression, is essential for p53-dependent growth suppression. A gene transfer strategy was used to demonstrate that under physiological conditions constitutive IMPD expression prevents p53-dependent growth suppression. In these studies, expression of bax and waf1, genes implicated in p53-dependent growth suppression in response to DNA damage, remains elevated in response to p53. These findings indicate that under physiological conditions IMPD is a rate-determining factor for p53-dependent growth regulation. In addition, they suggest that the impd gene may be epistatic to bax and waf1 in growth suppression. Because of the role of IMPD in the production and balance of GTP and ATP, essential nucleotides for signal transduction, these results suggest that p53 controls cell division signals by regulating purine ribonucleotide metabolism.

Poly(ADP-ribosyl)ation basally activated by DNA strand breaks reflects glutamate���nitric oxide neurotransmission

Poly(ADP-ribose) polymerase (PARP) transfers ADP ribose groups from NAD+ to nuclear proteins after activation by DNA strand breaks. PARP overactivation by massive DNA damage causes cell death via NAD+ and ATP depletion. Heretofore, PARP has been thought to be inactive under basal physiologic conditions. We now report high basal levels of PARP activity and DNA strand breaks in discrete neuronal populations of the brain, in ventricular ependymal and subependymal cells and in peripheral tissues. In some peripheral tissues, such as skeletal muscle, spleen, heart, and kidney, PARP activity is reduced only partially in mice with PARP-1 gene deletion (PARP-1���/���), implicating activity of alternative forms of PARP. Glutamate neurotransmission involving N-methyl-d-aspartate (NMDA) receptors and neuronal nitric oxide synthase (nNOS) activity in part mediates neuronal DNA strand breaks and PARP activity, which are diminished by NMDA antagonists and NOS inhibitors and also diminished in mice with targeted deletion of nNOS gene (nNOS���/���). An increase in NAD+ levels after treatment with NMDA antagonists or NOS inhibitors, as well as in nNOS���/��� mice, indicates that basal glutamate-PARP activity regulates neuronal energy dynamics.

Poly(ADP-ribose) polymerase gene disruption conferred mice resistant to streptozotocin-induced diabetes

Streptozotocin (STZ), a glucose analogue known to induce diabetes in experimental animals, causes DNA strand breaks and subsequent activation of poly(ADPribose) polymerase (Parp). Because Parp uses NAD as a substrate, extensive DNA damage will result in reduction of cellular NAD level. In fact, STZ induces NAD depletion and cell death in isolated pancreatic islets in vitro. Activation of Parp therefore is thought to play an important role in STZ-induced diabetes. In the present study, we established Parp-deficient (Parp���/���) mice by disrupting Parp exon 1 by using the homologous recombination technique. These mice were used to examine the possible involvement of Parp in STZ-induced ��-cell damage in vivo. The wild-type (Parp+/+) mice showed significant increases in blood glucose concentration from 129 mg/dl to 218, 370, 477, and 452 mg/dl on experimental days 1, 7, 21, and 60, respectively, after a single injection of 180 mg STZ/kg body weight. In contrast, the concentration of blood glucose in Parp���/��� mice remained normal up to day 7, slightly increased on day 21, but returned to normal levels on day 60. STZ injection caused extensive necrosis in the islets of Parp+/+ mice on day 1, with subsequent progressive islet atrophy and loss of functional �� cells from day 7. In contrast, the extent of islet ��-cell death and dysfunction was markedly less in Parp���/��� mice. Our findings clearly implicate Parp activation in islet ��-cell damage and glucose intolerance induced by STZ in vivo.

Crystal structures of bovine milk xanthine dehydrogenase and xanthine oxidase: Structure-based mechanism of conversion

Mammalian xanthine oxidoreductases, which catalyze the last two steps in the formation of urate, are synthesized as the dehydrogenase form xanthine dehydrogenase (XDH) but can be readily converted to the oxidase form xanthine oxidase (XO) by oxidation of sulfhydryl residues or by proteolysis. Here, we present the crystal structure of the dimeric (Mr, 290,000) bovine milk XDH at 2.1-��� resolution and XO at 2.5-��� resolution and describe the major changes that occur on the proteolytic transformation of XDH to the XO form. Each molecule is composed of an N-terminal 20-kDa domain containing two iron sulfur centers, a central 40-kDa flavin adenine dinucleotide domain, and a C-terminal 85-kDa molybdopterin-binding domain with the four redox centers aligned in an almost linear fashion. Cleavage of surface-exposed loops of XDH causes major structural rearrangement of another loop close to the flavin ring (Gln 423���Lys 433). This movement partially blocks access of the NAD substrate to the flavin adenine dinucleotide cofactor and changes the electrostatic environment of the active site, reflecting the switch of substrate specificity observed for the two forms of this enzyme.