Identificação de genes em Chromobacterium violaceum relacionados à resposta ao estresse

The sequencing of the genome of Chromobacterium violaceum identified one single circular chromosome of 4.8 Mb, in which approximately 40% of the founded ORFs are classified as hypothetical conserved or hypothetical. Some genic regions of biotechnological and biological interest had been characterized, e. g., environmental detoxification and DNA repair genes, respectively. Given this fact, the aim of this work was to identify genes of C. violaceum related to stress response, as the ones involved with mechanisms of DNA repair and/or genomic integrity maintenance. For this, a genomic library of C. violaceum was built in Escherichia coli strain DH10B (RecA-), in which clones were tested to UVC resistance, resulting in five candidates clones. In the PLH6A clone were identified four ORFs (CV_3721 to 3724). Two ORFs, CV_3722 and CV_3724, were subcloned and a synergic complementation activity was observed. The occurrence of an operon was confirmed using cDNA from C. violaceum in a RT-PCR assay. Further, it was observed the induction of the operon after the treatment with UVC. Thus, this operon was related to the stress response in C. violaceum. The mutagenesis assay with rifampicin after the treatment with UVC light showed high frequency of mutagenicity for the ORF CV_3722 (Pol III δ subunit). In this way, we propose that the C. violaceum δ subunit can act in DH10B in the translesion synthesis using Pol IV in a RecA independent-manner pathway. In growth curve assays other four clones (PLE1G, PLE7B, PLE10B and PLE12H) were able to complement the function at the dose 5 J/m2 and in mutagenicity assays PLE7B, PLE10B and PLE12H showed frequencies of mutation with significant differences upon the control (DH10B), demonstrating that in some way they are involved with the stress response in C. violaceum. These clones appear to be interrelated, probably regulated by a messenger molecule (eg., nucleotide c-di-GMP) and/or global regulatory molecule (eg., σS subunit of RNA polymerase).The results obtained contribute for a better genetic knowledge of this specie and its response mechanisms to environmental stress.

Proteômica comparativa de linhagens celulares humanas expostas a estresse oxidativo induzido por riboflavina fotossensibilizada

Reactive oxygen species (ROS) are continuously generated and can be derived from cellular metabolism or induced by exogenous factors, in addition, have the capacity to damage molecules like DNA and proteins. BER is considered the main route of DNA damage oxidative repair, however, several studies have demonstrated the importance of the proteins participation of other ways to correct these injuries. NER enzymes deficiency, such as CSB and XPC, acting in the damage recognition step in the two subways this system influences the effectiveness of oxidative damage repair. However, the mechanisms by which cells deficient in these enzymes respond to oxidative stress and its consequences still need to be better understood. Thus, the aim of this study was to perform a proteomic analysis of cell lines proficient and deficient in NER, exposed to oxidative stress, in order to identify proteins involved, directly or not, in response to oxidative stress and DNA repair. For this, three strains of human fibroblasts, MRC5-SV, CS1AN (CSBdeficient) and XP4PA (XPC-deficient) were treated with photosensitized riboflavin and then carried out the differentially expressed proteins identification by mass spectrometry. From the results, it was observed in MRC5-SV increase expression in most of the proteins involved in cellular defense, an expected response to a normal cell line subjected to stress. CS1AN showed a response disjointed, it is not possible to establish many interactions between the proteins identified, may be one explanation for their sensitivity to treatment with riboflavin and other oxidants and increased cell death probably by induction of pro-apoptotic pathways. Already XP4PA showed higher expression of apoptosis-blocking proteins, as there was inhibition or reduced expression of others involved with the activation of this process, suggesting the activation of an anti-apoptotic mechanism in this lineage, which may help explain the high susceptibility to develop cancers in XPC individuals. These results also contribute to elucidate action mechanisms of NER in oxidative damage and the understanding of important routes in the oxidative stress correlation, repair and malignant tumors formation

A Formação de Padres no Nordeste do Brasil (1894-1933)

The Nossa Senhora da Conceição Seminary, installed in 1894, by Dom Adauto Aurélio de Miranda Henriques, first Paraíba Bishop, and the Episcopal Seminary of the Sagrado Coração de Jesus, implanted in 1913, by Dom José Thomas Gomes da Silva, first Aracaju s Bishop diocese, were created as a result of lack of an official religious process proposed by the Brazilian Republic Proclamation, in 1889. With the appoint to enlarge the number of priests and change the image of the priest married and unrolled who used to identify the Catholic Church in the colonial and imperial Brazil. Such bishops developed into intellectuals in the government, dioceses and formation priest houses. I take as a study object, for this doctorate paper, the academic formation and priesthood developed in theses seminaries, from 1894 to 1933, once 1894 the year of João Pessoa Creation Seminar that was implied the Minor Course (preparation) and the Major one (built by Philosophy and God related studies) and the research limit year of 1933, is concerned about the Major Sergipe Seminary ending, which was created and has worked offering the Minor and Major courses, from 1913 to 1933. Showing the teaching models that guided and leaded the priest formation, referred as Seminaries, and the application result is the objective of this investigation. To comprehend the teaching models seminaries studied, my research line is the Catholic Church theme and priest formation in Brazil. In front of the object and the objective desired, I chose the historical comparative method and the scholars modals notions of Araujo de Barros (2004) and the Sirinelli intellectuals (1996). Such references allowed me to analyze the formation given in the seminary and seminarian participation and actions, included the sequence after the scholars formation. The thesis defended is that the teaching model developed in the Brazilian Seminaries, created after a non official religious process in the Brazilian government, deal with a model of one unique center (Seminary formation and aim pre arranged by Santa Sé), although adapted, presuming the local reality and formation structure (privileged not only spiritual and moral speaking, but intellectual also), was it responsible for intellectuals generations (teachers priests, educationalist priest, journalists priests and so on) that boost the education in Brazil. During the Republic first three decades, when, in thesis, the Government was becoming free religion, i.e., the government did not subsidize the Church anymore, and the Government, among others aspects, did not received any Church care to help the public teaching in the country. The investigation reveled accede, by bishops and their followers, such as by the Concílio de Trento pre concept, or by the others ideas, leading by the priests formation in Seminaries. By creating and stalling diocese Seminary, Bishop Dom Adauto and Dom José went further their functions, by the time they built inside themselves a teaching model thought from the main pedagogic logic, based on several religious exercises, moral and ethic, considered by themselves several knowledge connected to humanity, philosophy and God related studies). Following clearly rationalism principle (the way of teaching, which each subject has its own teacher and this class get together students with the same knowledge, regardless of age) and efficiency (trying to teach the whole content in each class), the Seminaries researched developed a whole education, allowed the structure of a spiritual education, moral and intellectual, for a quality developed by priests, including different levels that they used to performance. Their bottom line, actions and priest matter achievement allowed their broad fulfillment, in the way that priests matter were associated with cultural, educational, welfare assistance, at last, intellectuals

Identificação de genes em Chromobacterium violaceum relacionados à resposta ao estresse

The sequencing of the genome of Chromobacterium violaceum identified one single circular chromosome of 4.8 Mb, in which approximately 40% of the founded ORFs are classified as hypothetical conserved or hypothetical. Some genic regions of biotechnological and biological interest had been characterized, e. g., environmental detoxification and DNA repair genes, respectively. Given this fact, the aim of this work was to identify genes of C. violaceum related to stress response, as the ones involved with mechanisms of DNA repair and/or genomic integrity maintenance. For this, a genomic library of C. violaceum was built in Escherichia coli strain DH10B (RecA-), in which clones were tested to UVC resistance, resulting in five candidates clones. In the PLH6A clone were identified four ORFs (CV_3721 to 3724). Two ORFs, CV_3722 and CV_3724, were subcloned and a synergic complementation activity was observed. The occurrence of an operon was confirmed using cDNA from C. violaceum in a RT-PCR assay. Further, it was observed the induction of the operon after the treatment with UVC. Thus, this operon was related to the stress response in C. violaceum. The mutagenesis assay with rifampicin after the treatment with UVC light showed high frequency of mutagenicity for the ORF CV_3722 (Pol III δ subunit). In this way, we propose that the C. violaceum δ subunit can act in DH10B in the translesion synthesis using Pol IV in a RecA independent-manner pathway. In growth curve assays other four clones (PLE1G, PLE7B, PLE10B and PLE12H) were able to complement the function at the dose 5 J/m2 and in mutagenicity assays PLE7B, PLE10B and PLE12H showed frequencies of mutation with significant differences upon the control (DH10B), demonstrating that in some way they are involved with the stress response in C. violaceum. These clones appear to be interrelated, probably regulated by a messenger molecule (eg., nucleotide c-di-GMP) and/or global regulatory molecule (eg., σS subunit of RNA polymerase).The results obtained contribute for a better genetic knowledge of this specie and its response mechanisms to environmental stress.