The Atp-dependent Rna Helicase Hrpb Plays An Important Role In Motility And Biofilm Formation In Xanthomonas Citri Subsp Citri

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)RNA helicases are enzymes that catalyze the separation of double-stranded RNA (dsRNA) using the free energy of ATP binding and hydrolysis. DEAD/DEAH families participate in many different aspects of RNA metabolism, including RNA synthesis, RNA folding, RNA-RNA interactions, RNA localization and RNA degradation. Several important bacterial DEAD/DEAH-box RNA helicases have been extensively studied. In this study, we characterize the ATP-dependent RNA helicase encoded by the hrpB (XAC0293) gene using deletion and genetic complementation assays. We provide insights into the function of the hrpB gene in Xanthomonas citri subsp. citri by investigating the roles of hrpB in biofilm formation on abiotic surfaces and host leaves, cell motility, host virulence of the citrus canker bacterium and growth in planta. Results: The hrpB gene is highly conserved in the sequenced strains of Xanthomonas. Mutation of the hrpB gene (Delta hrpB) resulted in a significant reduction in biofilms on abiotic surfaces and host leaves. Delta hrpB also exhibited increased cell dispersion on solid medium plates. Delta hrpB showed reduced adhesion on biotic and abiotic surfaces and delayed development in disease symptoms when sprayed on susceptible citrus leaves. Quantitative reverse transcription-PCR assays indicated that deletion of hrpB reduced the expression of four type IV pili genes. The transcriptional start site of fimA (XAC3241) was determined using rapid amplification of 5'-cDNA Ends (5' RACE). Based on the results of fimA mRNA structure predictions, the fimA 5' UTR may contain three different loops. HrpB may be involved in alterations to the structure of fimA mRNA that promote the stability of fimA RNA. Conclusions: Our data show that hrpB is involved in adherence of Xanthomonas citri subsp. citri to different surfaces. In addition, to the best of our knowledge, this is the first time that a DEAH RNA helicase has been implicated in the regulation of type IV pili in Xanthomonas.1635INCT CitrusCAPES/PSDE fellowship [99999.002657/2014-07]CNPqCoordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq

History and social qualitative research : reflections on comparative history and life history

Este ensayo teórico tiene el objetivo de reflexionar, desde una perspectiva brasileña, sobre el uso del método histórico en las ciencias sociales en general, y en las áreas de los estudios organizacionales y de la administración pública en particular. Para ello, con base en una revisión bibliográfica, se eligió profundizar la reflexión a partir de los métodos histórico-comparativos y de la historia de vida, dentro del paradigma cualitativo, con el fin de ofrecer una perspectiva más especializada sobre las implicancias de tales métodos para las subdisciplinas antes mencionadas. Se concluye que el método histórico en general, así como el histórico-comparativo y la historia de vida en particular, promueven nuevas formas de comprensión de la realidad, que conectan lo singular con lo global, lo objetivo con lo subjetivo, lo pasado con lo contemporáneo, generando perspectivas innovadoras para el análisis social, organizacional y político.This theoretical essay aims to reflect, in a Brazilian perspective, on the use of the historical method in the social sciences in general, and in the areas of organizational studies and public administration. To do this, based on a bibliographical review, we have chosen to deepen the reflection from the historical-comparative method and life history, within the qualitative paradigm, in order to offer a more specialized perspective on the implications of such methods for the aforementioned subdisciplines. We conclude that the historical method in general, as well as the historical-comparative method and life history method, in particular, promote new forms of understanding reality, by connecting the singular, the global, the objective with the subjective, the past with the contemporary, and generating innovative perspectives for social, organizational and political analyses

Ligand-based chemoinformatic discovery of a novel small molecule inhibitor targeting CDC25 dual specificity phosphatases and displaying in vitro efficacy against melanoma cells

CDC25 phosphatases are important regulators of the cell cycle and represent promising targets for anticancer drug discovery. We recently identified NSC 119915 as a new quinonoid CDC25 inhibitor with potent anticancer activity. In order to discover more active analogs of NSC 119915, we performed a range of ligand-based chemoinformatic methods against the full ZINC drug-like subset and the NCI lead-like set. Nine compounds (3, 5?9, 21, 24, and 25) were identified with Ki values for CDC25A, -B and -C ranging from 0.01 to 4.4 ?M. One of these analogs, 7, showed a high antiproliferative effect on human melanoma cell lines, A2058 and SAN. Compound 7 arrested melanoma cells in G2/M, causing a reduction of the protein levels of CDC25A and, more consistently, of CDC25C. Furthermore, an intrinsic apoptotic pathway was induced, which was mediated by ROS, because it was reverted in the presence of antioxidant N-acetyl-cysteine (NAC). Finally, 7 decreased the protein levels of phosphorylated Akt and increased those of p53, thus contributing to the regulation of chemosensitivity through the control of downstream Akt pathways in melanoma cells. Taken together, our data emphasize that CDC25 could be considered as a possible oncotarget in melanoma cells and that compound 7 is a small molecule CDC25 inhibitor that merits to be further evaluated as a chemotherapeutic agent for melanoma, likely in combination with other therapeutic compounds

Bradykinin B2 Receptors of Dendritic Cells, Acting as Sensors of Kinins Proteolytically Released by Trypanosoma cruzi, Are Critical for the Development of Protective Type-1 Responses

Although the concept that dendritic cells (DCs) recognize pathogens through the engagement of Toll-like receptors is widely accepted, we recently suggested that immature DCs might sense kinin-releasing strains of Trypanosoma cruzi through the triggering of G-protein-coupled bradykinin B2 receptors (B2R). Here we report that C57BL/6.B2R−/− mice infected intraperitoneally with T. cruzi display higher parasitemia and mortality rates as compared to B2R+/+ mice. qRT-PCR revealed a 5-fold increase in T. cruzi DNA (14 d post-infection [p.i.]) in B2R−/− heart, while spleen parasitism was negligible in both mice strains. Analysis of recall responses (14 d p.i.) showed high and comparable frequencies of IFN-γ-producing CD4+ and CD8+ T cells in the spleen of B2R−/− and wild-type mice. However, production of IFN-γ by effector T cells isolated from B2R−/− heart was significantly reduced as compared with wild-type mice. As the infection continued, wild-type mice presented IFN-γ-producing (CD4+CD44+ and CD8+CD44+) T cells both in the spleen and heart while B2R−/− mice showed negligible frequencies of such activated T cells. Furthermore, the collapse of type-1 immune responses in B2R−/− mice was linked to upregulated secretion of IL-17 and TNF-α by antigen-responsive CD4+ T cells. In vitro analysis of tissue culture trypomastigote interaction with splenic CD11c+ DCs indicated that DC maturation (IL-12, CD40, and CD86) is controlled by the kinin/B2R pathway. Further, systemic injection of trypomastigotes induced IL-12 production by CD11c+ DCs isolated from B2R+/+ spleen, but not by DCs from B2R−/− mice. Notably, adoptive transfer of B2R+/+ CD11c+ DCs (intravenously) into B2R−/− mice rendered them resistant to acute challenge, rescued development of type-1 immunity, and repressed TH17 responses. Collectively, our results demonstrate that activation of B2R, a DC sensor of endogenous maturation signals, is critically required for development of acquired resistance to T. cruzi infection

Antagonizing S1P3 Receptor with Cell-Penetrating Pepducins in Skeletal Muscle Fibrosis

S1P is the final product of sphingolipid metabolism, which interacts with five widely expressed GPCRs (S1P1-5). Increasing numbers of studies have indicated the importance of S1P3 in various pathophysiological processes. Recently, we have identified a pepducin (compound KRX-725-II) acting as an S1P3 receptor antagonist. Here, aiming to optimize the activity and selectivity profile of the described compound, we have synthesized a series of derivatives in which Tyr, in position 4, has been substituted with several natural aromatic and unnatural aromatic and non-aromatic amino acids. All the compounds were evaluated for their ability to inhibit vascular relaxation induced by KRX-725 (as S1P3 selective pepducin agonist) and KRX-722 (an S1P1-selective pepducin agonist). Those selective towards S1P3 (compounds V and VII) were also evaluated for their ability to inhibit skeletal muscle fibrosis. Finally, molecular dynamics simulations were performed to derive information on the preferred conformations of selective and unselective antagonists

TLR9 Signaling Suppresses the Canonical Plasma Cell Differentiation Program in Follicular B Cells

The relative potency and quality of mouse B cell response to Toll-like receptors (TLRs) signaling varies significantly depending on the B cell subset and on the TLR member being engaged. Although it has been shown that marginal zone cells respond faster than follicular (FO) splenic B cells to TLR4 stimulus, FO B cells retain full capacity to proliferate and generate plasmablasts and plasma cells (PBs/PCs) with 2–3 days delayed kinetics. It is not clear whether this scenario could be extended to other members of the TLR family. Here, using quantitative cell culture conditions optimized for B cell growth and differentiation, we show that TLR9 signaling by CpG, while promoting vigorous proliferation, completely fails to induce differentiation of FO B cells into PBs/PCs. Little or absent Ig secretion following TLR9 stimulus was accompanied by lack of expression of cell surface markers and canonical transcription factors involved in PB/PC differentiation. Moreover, not only TLR9 did not induce plasmocyte differentiation, but it also strongly inhibited the massive PB/PC differentiation of FO B cells triggered by LPS/TLR4. Our study reveals unexpected opposite roles for TLR4 and TLR9 in the control of plasma cell differentiation program and disagrees with previous conclusions obtained in high-density cultures conditions on the generation of plasmocytes by TRL9 signaling. The potential implications of these findings on the role of TLR9 in controlling self-tolerance, clonal sizes and regulation of humoral responses are discussed

History and social qualitative research : reflections on comparative history and life history

Este ensayo teórico tiene el objetivo de reflexionar, desde una perspectiva brasileña, sobre el uso del método histórico en las ciencias sociales en general, y en las áreas de los estudios organizacionales y de la administración pública en particular. Para ello, con base en una revisión bibliográfica, se eligió profundizar la reflexión a partir de los métodos histórico-comparativos y de la historia de vida, dentro del paradigma cualitativo, con el fin de ofrecer una perspectiva más especializada sobre las implicancias de tales métodos para las subdisciplinas antes mencionadas. Se concluye que el método histórico en general, así como el histórico-comparativo y la historia de vida en particular, promueven nuevas formas de comprensión de la realidad, que conectan lo singular con lo global, lo objetivo con lo subjetivo, lo pasado con lo contemporáneo, generando perspectivas innovadoras para el análisis social, organizacional y político.This theoretical essay aims to reflect, in a Brazilian perspective, on the use of the historical method in the social sciences in general, and in the areas of organizational studies and public administration. To do this, based on a bibliographical review, we have chosen to deepen the reflection from the historical-comparative method and life history, within the qualitative paradigm, in order to offer a more specialized perspective on the implications of such methods for the aforementioned subdisciplines. We conclude that the historical method in general, as well as the historical-comparative method and life history method, in particular, promote new forms of understanding reality, by connecting the singular, the global, the objective with the subjective, the past with the contemporary, and generating innovative perspectives for social, organizational and political analyses