SpoIIE governs the phosphorylation state of a protein regulating transcription factor sigma F during sporulation in Bacillus subtilis.

Cell-specific activation of the transcription factor sigma F during sporulation in Bacillus subtilis is controlled by a regulatory pathway involving the proteins SpoIIE, SpoIIAA, and SpoIIAB. SpoIIAB is an antagonist of sigma F, and SpoIIAA, which is capable of overcoming SpoIIAB-mediated inhibition of sigma F, is an antagonist of SpoIIAB. SpoIIAA is, in turn, negatively regulated by SpoIIAB, which phosphorylates SpoIIAA on serine 58. SpoIIAA is also positively regulated by SpoIIE, which dephosphorylates SpoIIAA-P, the phosphorylated form of SpoIIAA. Here, isoelectric focusing and Western blot analysis were used to examine the phosphorylation state of SpoIIAA in vivo. SpoIIAA was found to be largely in the phosphorylated state during sporulation in wild-type cells but a significant portion of the protein that was unphosphorylated could also be detected. Consistent with the idea that SpoIIE governs dephosphorylation of SpoIIAA-P, SpoIIAA was entirely in the phosphorylated state in spoIIE mutant cells. Conversely, overexpression of spoIIE led to an increase in the ratio of unphosphorylated SpoIIAA to SpoIIAA-P and caused inappropriate activation of sigma F in the predivisional sporangium. We also show that a mutant form of SpoIIAA (SpoIIAA-S58T) in which serine 58 was replaced with threonine was present exclusively as SpoIIAA-P, a finding that confirms previous biochemical evidence that the mutant protein is an effective substrate for the SpoIIAB kinase but that SpoIIAA-S58T-P cannot be dephosphorylated by SpoIIE. We conclude that SpoIIE plays a crucial role in controlling the phosphorylation state of SpoIIAA during sporulation and thus in governing the cell-specific activation of sigma F.

Crystal structure of the T state of allosteric yeast chorismate mutase and comparison with the R state.

The crystal structure of the tyrosine-bound T state of allosteric yeast Saccharomyces cerevisiae chorismate mutase was solved by molecular replacement at a resolution of 2.8 angstroms using a monomer of the R-state structure as the search model. The allosteric inhibitor tyrosine was found to bind in the T state at the same binding site as the allosteric activator tryptophan binds in the R state, thus defining one regulatory binding site for each monomer. Activation by tryptophan is caused by the larger steric size of its side chain, thereby pushing apart the allosteric domain of one monomer and helix H8 of the catalytic domain of the other monomer. Inhibition is caused by polar contacts of tyrosine with Arg-75 and Arg-76 of one monomer and with Gly-141, Ser-142, and Thr-145 of the other monomer, thereby bringing the allosteric and catalytic domains closer together. The allosteric transition includes an 8 degree rotation of each of the two catalytic domains relative to the allosteric domains of each monomer (domain closure). Alternatively, this transition can be described as a 15 degree rotation of the catalytic domains of the dimer relative to each other.

Oral tolerance in myelin basic protein T-cell receptor transgenic mice: suppression of autoimmune encephalomyelitis and dose-dependent induction of regulatory cells.

Orally administered antigens induce a state of immunologic hyporesponsiveness termed oral tolerance. Different mechanisms are involved in mediating oral tolerance depending on the dose fed. Low doses of antigen generate cytokine-secreting regulatory cells, whereas high doses induce anergy or deletion. We used mice transgenic for a T-cell receptor (TCR) derived from an encephalitogenic T-cell clone specific for the acetylated N-terminal peptide of myelin basic protein (MBP) Ac-1-11 plus I-Au to test whether a regulatory T cell could be generated from the same precursor cell as that of an encephalitogenic Th1 cell and whether the induction was dose dependent. The MBP TCR transgenic mice primarily have T cells of a precursor phenotype that produce interleukin 2 (IL-2) with little interferon gamma (IFN-gamma), IL-4, or transforming growth factor beta (TGF-beta). We fed transgenic animals a low-dose (1 mg x 5) or high-dose (25 mg x 1) regimen of mouse MBP and without further immunization spleen cells were tested for cytokine production. Low-dose feeding induced prominent secretion of IL-4, IL-10, and TGF-beta, whereas minimal secretion of these cytokines was observed with high-dose feeding. Little or no change was seen in proliferation or IL-2/IFN-gamma secretion in fed animals irrespective of the dose. To demonstrate in vivo functional activity of the cytokine-secreting cells generated by oral antigen, spleen cells from low-dose-fed animals were adoptively transferred into naive (PLJ x SJL)F1 mice that were then immunized for the development of experimental autoimmune encephalomyelitis (EAE). Marked suppression of EAE was observed when T cells were transferred from MBP-fed transgenic animals but not from animals that were not fed. In contrast to oral tolerization, s.c. immunization of transgenic animals with MBP in complete Freund's adjuvant induced IFN-gamma-secreting Th1 cells in vitro and experimental encephalomyelitis in vivo. Despite the large number of cells reactive to MBP in the transgenic animals, EAE was also suppressed by low-dose feeding of MBP prior to immunization. These results demonstrate that MBP-specific T cells can differentiate in vivo into encephalitogenic or regulatory T cells depending upon the context by which they are exposed to antigen.

Identification of a member of the interferon regulatory factor family that binds to the interferon-stimulated response element and activates expression of interferon-induced genes.

A family of interferon (IFN) regulatory factors (IRFs) have been shown to play a role in transcription of IFN genes as well as IFN-stimulated genes. We report the identification of a member of the IRF family which we have named IRF-3. The IRF-3 gene is present in a single copy in human genomic DNA. It is expressed constitutively in a variety of tissues and no increase in the relative steady-state levels of IRF-3 mRNA was observed in virus-infected or IFN-treated cells. The IRF-3 gene encodes a 50-kDa protein that binds specifically to the IFN-stimulated response element (ISRE) but not to the IRF-1 binding site PRD-I. Overexpression of IRF-3 stimulates expression of the IFN-stimulated gene 15 (ISG15) promoter, an ISRE-containing promoter. The murine IFNA4 promoter, which can be induced by IRF-1 or viral infection, is not induced by IRF-3. Expression of IRF-3 as a Gal4 fusion protein does not activate expression of a chloramphenicol acetyltransferase reporter gene containing repeats of the Gal4 binding sites, indicating that this protein does not contain the transcription transactivation domain. The high amino acid homology between IRF-3 and ISG factor 3 gamma polypeptide (ISGF3 gamma) and their similar binding properties indicate that, like ISGF3 gamma, IRF-3 may activate transcription by complex formation with other transcriptional factors, possibly members of the Stat family. Identification of this ISRE-binding protein may help us to understand the specificity in the various Stat pathways.

Opposing early and late effects of insulin-like growth factor I on differentiation and the cell cycle regulatory retinoblastoma protein in skeletal myoblasts.

The mechanisms by which insulin-like growth factors (IGFs) can be both mitogenic and differentiation-promoting in skeletal myoblasts are unclear because these two processes are believed to be mutually exclusive in this tissue. The phosphorylation state of the ubiquitous nuclear retinoblastoma protein (Rb) plays an important role in determining whether myoblasts proliferate or differentiate: Phosphorylated Rb promotes mitogenesis, whereas un- (or hypo-) phosphorylated Rb promotes cell cycle exit and differentiation. We hypothesized that IGFs might affect the fate of myoblasts by regulating the phosphorylation of Rb. Although long-term IGF treatment is known to stimulate differentiation, we find that IGFs act initially to inhibit differentiation and are exclusively mitogenic. These early effects of IGFs are associated with maintenance of Rb phosphorylation typical of proliferating cells; upregulation of the gene expression of cyclin-dependent kinase 4 and cyclin D1, components of a holoenzyme that plays a principal role in mediating Rb phosphorylation; and marked inhibition of the gene expression of myogenin, a member of the MyoD family of skeletal muscle-specific transcription factors that is essential in muscle differentiation. We also find that IGF-induced inhibition of differentiation occurs through a process that is independent of its mitogenic effects. We demonstrate, thus, that IGFs regulate Rb phosphorylation and cyclin D1 and cyclin-dependent kinase 4 gene expression; together with their biphasic effects on myogenin expression, these results suggest a mechanism by which IGFs are initially mitogenic and subsequently differentiation-promoting in skeletal muscle.

A qualidade da regulação estatal no Brasil: uma análise a partir de indicadores de qualidade

As primeiras agências reguladoras foram criadas a partir da segunda metade dos anos 1990, e a mais recente delas, em 2005. Com as agências surgiram também os atores privados regulados, os usuários e consumidores, e uma nova forma de interação entre os Poderes Executivo, Legislativo e Judiciário. Esses atores participam e dão forma ao processo de aprendizagem institucional das agências. Passado o período de criação e após quase duas décadas de existência, é necessária uma visão crítica sobre as agências. Propõe-se, então, um método de avaliação regulatória a partir de três variáveis que serão decompostas em diversas subvariáveis (quesitos a serem respondidos objetivamente). A primeira variável, institucionalização, mede as regras aplicáveis à própria agência: características dos mandatos dos dirigentes, autonomia decisória, autonomia financeira e de gestão de pessoal. A segunda, procedimentalização, ocupa-se do processo de tomada de decisão da agência e de sua transparência. Ambas as variáveis procuram medir as agências do ponto de vista formal, a partir de normas aplicáveis (leis, decretos, resoluções, portarias etc.), e pela prática regulatória, com base nos fatos ocorridos demonstrados por meio de documentos oficiais (decretos de nomeação, decisões, relatórios de atividade das próprias agências etc.). A última variável, judicialização, aponta as várias vezes em que a decisão administrativa muda de status e o nível de confirmação dessas decisões pelo Poder Judiciário. O modelo teórico de avaliação das agências ora apresentado é aplicado e testado em três setores que são submetidos à regulação econômica e contam com forte presença de atores sociais e empresa estatal federal. Assim, as agências analisadas foram: Agência Nacional de Energia Elétrica ANEEL, Agência Nacional de Telecomunicações ANATEL e Agência Nacional de Aviação Civil ANAC. Em termos gerais, não é possível garantir a existência de um isoformismo entre essas agências, nem mesmo entre agências criadas em momentos diferentes e por presidentes distintos. Também não foi possível demonstrar que a interferência política seja uma marca de um único governo. A ANATEL, a melhor avaliada das três agências, destaca-se pelo rigor de suas normas que seu processo decisório reflete. A ANEEL e a ANAC tiveram uma avaliação mediana já que apresentaram avaliação sofrível quanto ao processo, mas mostraram ter instituições (regras) um pouco melhores.

Impacto socioeconômico das políticas sanitárias sobre os estabelecimentos avícolas comerciais de postura da regional agropecuária de Limeira, estado de São Paulo

É reconhecida a importância da avicultura para o agronegócio e para o desenvolvimento do Brasil. Mas o País tem enfrentado grande concorrência internacional, que se traduz em barreiras sanitárias e exigências cada vez maiores de controle de seu rebanho por parte dos importadores. Neste sentido, o Ministério da Agricultura, Pecuária e Abastecimento (MAPA) publicou uma série de atos legais para viabilizar a organização dos programas de saúde animal, entre eles as Instruções Normativas n° 56/2007, n° 59/2009, n°36/2012 e n°10/2013, para estabelecer os procedimentos para registro e fiscalização de estabelecimentos avícolas comerciais e de reprodução, voltados à biosseguridade do sistema avícola. Como todos os setores produtivos, a avicultura é fortemente influenciada pela sua estrutura de custos, de modo que há um certo sentimento de que a adequação às medidas de biosseguridade, preconizadas pelas normativas, pode impactar a atividade de produção de ovos comerciais de modo a levar, eventualmente, alguns avicultores ao abandono da mesma. O objetivo deste trabalho foi analisar o impacto socioeconômico das políticas sanitárias para estabelecimentos avícolas comerciais de postura da regional agropecuária de Limeira, Estado de São Paulo, analisando o perfil socioeconômico da propriedade e do produtor e, estimando o custo de produção e seu impacto em atendimento às medidas de biosseguridade preconizadas pelas normativas. Para a análise do perfil das propriedades foram utilizados os dados dos 28 estabelecimentos avícolas, levantados por meio de documentação para registro. Para a análise do perfil do produtor e para estimar os custos de produção foram considerados dez estabelecimentos, voluntários à pesquisa, visitados entre os meses de junho e julho de 2013. O resultado do estudo sugere que as adequações à biosseguridade podem ser factíveis de serem realizadas economicamente, sendo o custo relativamente pequeno frente aos possíveis riscos de enfermidades, representando entre 1,61% e 2,09% do custo total de produção. No entanto, as sucessivas alterações nas legislações podem fazer com que o programa de sanidade avícola perca a credibilidade diante da sociedade e dos produtores que demonstram resistência às mudanças nos paradigmas zoosanitários

A Regulatory Compliance Plan for Decommissioning a Sodium Phosphate Manufacturing Plant

Sodium phosphates are a class of chemicals that have been widely employed in commercial and consumer applications. However, declining use of these chemicals due to environmental concerns has lead to restructuring within the industry that has caused, and is likely to continue to cause, reduction of sodium phosphate production capacity. Closure of a sodium phosphate manufacturing plant necessitates decommissioning and decontamination activities that are subject to a variety of federal, state, and local regulations. A compliance plan was developed to provide a blueprint for ensuring that all federal regulatory requirements are met, however, site dependent state and local requirements were excluded. The compliance plan provides a framework that addresses project team formation and project planning, regulatory requirements, identification of affected processing equipment, plant pre-shutdown activities, waste stream identification and waste management facilities, safety, training, and emergency preparedness planning, and project decommissioning remedial actions. This regulatory compliance plan will enable sodium phosphate plant operators to complete decontamination and decommissioning work in a timely, efficient, compliant, and cost effective manner.

The EU’s online gambling regulatory approach and the crisis of legal modernity. EU Centre Singapore Working Paper No. 19, January 2014

Online gambling is a fast growing service activity in the world - its economic significance is clearly shown by the high level of innovation by gambling operators all over the world, as well as by the increasing amount of tax revenues generated in those States that allow this activity. Nevertheless, states face many difficulties in controlling and regulating online gambling, given the specific nature of the Internet, and the never-ending quest by gamblers for new gaming websites that offer superior odds, a wider gaming variety, and greater bets combination. In this working paper, Dr Salvatore Casabona examines the legality of online gambling in the context of the European Union (EU), and discusses the Union's regulatory approach to online gambling, the lack of harmonisation and the issue of member state sovereignty at the crossroad of European Law on online gambling, and the potential for a new regulatory paradigm to emerge.

Nuclear power industry : joint hearing before the Committee on Energy and Natural Resources and the Subcommittee on Energy and Water Development of the Committee on Appropriations, United States Senate, One Hundred Seventh Congress, first session to conduct oversight on the state of the nuclear power industry and the future of the industry in a comprehensive energy strategy, May 3, 2001.

Shipping list no.: 2001-0300-P.

Regulatory water quality monitoring networks : statistical and economic considerations /

Mode of access: Internet.

State solar energy legislation of 1977 : a review of statutes relating to buildings /

Feb. 1979.

Annual report, Transportation Regulatory Fund, FY ...

Description based on; FY 1988; title from cover.

Illinois Department of Agriculture regulatory supplement.

"An addendum to the Annual report, fiscal year ..., July 1, ... - June 30, ..."

Potentially infectious medical waste : a summary of regulatory requirements : general requirements.

Caption title.