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Análise genética e funcional do gene rnr, uma exoribonuclease de Burkholderia cenocepacia: implicações na resposta ao stresse térmico, na regulação do ciclo celular e na virulência
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Algumas espécies bacterianas pertencentes ao Complexo Burkholderia cepacia (Bcc) são patogénicos oportunistas de humanos com o sistema imunitário comprometido e em especial de pacientes com fibrose quística (FQ). O ambiente pulmonar destes doentes permite a estas bactérias uma colonização eficaz criando infecções capazes de conduzir à morte de pacientes. A variedade de potenciais factores e mecanismos de virulência destes organismos têm sido objecto de vários estudos. Neste trabalho avaliamos o papel da RNase R, uma exoribonuclease 3’-5’, na virulência de Burkholderia cenocepacia K56-2, uma espécie pertencente à linhagem epidémica de Edimburgo-Toronto (ET)-12. Este microrganismo apresenta uma elevada capacidade de resposta a alterações das condições ambientais e a remodelação da sua expressão genética é um processo fundamental nesta resposta. Neste sentido o estudo da maquinaria de degradação do RNA e principalmente as RNases é fundamental. A RNase R não só tem vindo a ser implicada na virulência em vários organismos patogénicos para o Homem como também está descrita como uma importante proteína de resposta a stresse. Neste trabalho é demonstrado um papel fundamental da RNase R para o funcionamento correcto do ciclo celular. A ausência desta proteína leva a septação incompleta das células e diminui a viabilidade celular neste microrganismo. Comprova-se ainda, in vivo no modelo Galleria mellonella, que a proteína RNase R é importante na virulência de B.cenocepacia e que a expressão deste gene aparece aumentada em stresses ambientais como por exemplo a resposta ao choque do frio. Este estudo é o primeiro efectuado acerca do papel das RNases, em particular a RNase R, no metabolismo e virulência de B. cenocepacia. Demonstramos a importância desta proteína, principalmente na virulência e na divisão celular e por isso deverá ser de futuro uma proteína de eleição para estudos mais detalhados em Burkholderia e noutros organismos patogénicos.
Some of the members of Burkholdeira cepacia complex (Bcc) are opportunistic pathogens to immunocompromised patients, especially those with Cystic Fibrosis. The specific environment existent in the lungs of these patients allows an efficient bacterial colonization, giving rise to infections capable of leading to the death of the human host. The variety of potential factors and virulence mechanisms in these species have been highly explored. In this work, the role of RNase R, an exoribonuclease 3’-5’, in the virulence of Burkholderia cenocepacia K56-2 belonging to the epidemic lineage Edinburgh-Toronto (ET-12) is assessed. This microorganism has a great ability to quickly adapt to environmental changes and the remodel of their genetic expression profiles is a fundamental step in this response. Therefore, the study of the cellular machinery for RNA degradation, with RNases at the top, is fundamental to understand these mechanisms. RNase R not only has already been associated with virulence in human pathogens but has also been described as an important protein in stress responses. In this work, it is shown the role of RNase R for a proper progress of the bacteria cell cycle. The absence of this protein leads to an incomplete septation and a decrease in the cell viability in this microorganism. It is also show, in vivo in the Galleria mellonella model, that RNase R is important to the virulence of B. cenocepacia K56-2 and also that the expression of the encoding gene is increased in response to environmental stresse, as an example to the cold shock response. This study is the first regarding RNases, particularly RNase R, and their influence in the metabolism and virulence of B. cenocepacia. It is demonstrated here the importance of this protein in the cell division and virulence factor and it should be seen as another selected target for future studies in Burkholderia and another pathogenic microorganisms.
Some of the members of Burkholdeira cepacia complex (Bcc) are opportunistic pathogens to immunocompromised patients, especially those with Cystic Fibrosis. The specific environment existent in the lungs of these patients allows an efficient bacterial colonization, giving rise to infections capable of leading to the death of the human host. The variety of potential factors and virulence mechanisms in these species have been highly explored. In this work, the role of RNase R, an exoribonuclease 3’-5’, in the virulence of Burkholderia cenocepacia K56-2 belonging to the epidemic lineage Edinburgh-Toronto (ET-12) is assessed. This microorganism has a great ability to quickly adapt to environmental changes and the remodel of their genetic expression profiles is a fundamental step in this response. Therefore, the study of the cellular machinery for RNA degradation, with RNases at the top, is fundamental to understand these mechanisms. RNase R not only has already been associated with virulence in human pathogens but has also been described as an important protein in stress responses. In this work, it is shown the role of RNase R for a proper progress of the bacteria cell cycle. The absence of this protein leads to an incomplete septation and a decrease in the cell viability in this microorganism. It is also show, in vivo in the Galleria mellonella model, that RNase R is important to the virulence of B. cenocepacia K56-2 and also that the expression of the encoding gene is increased in response to environmental stresse, as an example to the cold shock response. This study is the first regarding RNases, particularly RNase R, and their influence in the metabolism and virulence of B. cenocepacia. It is demonstrated here the importance of this protein in the cell division and virulence factor and it should be seen as another selected target for future studies in Burkholderia and another pathogenic microorganisms.
Descrição
Tese de mestrado. Biologia (Biologia Humana e Ambiente). Universidade de Lisboa, Faculdade de Ciências, 2010
Palavras-chave
Biologia celular Fibrose quística Resistência aos antibióticos Expressão genética Teses de mestrado - 2010