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Mycobacterium tuberculosis nucleoside diphosphate kinase inactivates small GTPases leading to evasion of innate immunity.
Sun, Jim; Singh, Vijender; Lau, Alice; Stokes, Richard W; Obregón-Henao, Andrés; Orme, Ian M; Wong, Dennis; Av-Gay, Yossef; Hmama, Zakaria.
Afiliação
  • Sun J; Division of Infectious Diseases, Department of Medicine and Vancouver Coastal Health Research Institute, University of British Columbia, Vancouver, British Columbia, Canada.
PLoS Pathog ; 9(7): e1003499, 2013.
Article em En | MEDLINE | ID: mdl-23874203
Defining the mechanisms of Mycobacterium tuberculosis (Mtb) persistence in the host macrophage and identifying mycobacterial factors responsible for it are keys to better understand tuberculosis pathogenesis. The emerging picture from ongoing studies of macrophage deactivation by Mtb suggests that ingested bacilli secrete various virulence determinants that alter phagosome biogenesis, leading to arrest of Mtb vacuole interaction with late endosomes and lysosomes. While most studies focused on Mtb interference with various regulators of the endosomal compartment, little attention was paid to mechanisms by which Mtb neutralizes early macrophage responses such as the NADPH oxidase (NOX2) dependent oxidative burst. Here we applied an antisense strategy to knock down Mtb nucleoside diphosphate kinase (Ndk) and obtained a stable mutant (Mtb Ndk-AS) that displayed attenuated intracellular survival along with reduced persistence in the lungs of infected mice. At the molecular level, pull-down experiments showed that Ndk binds to and inactivates the small GTPase Rac1 in the macrophage. This resulted in the exclusion of the Rac1 binding partner p67(phox) from phagosomes containing Mtb or Ndk-coated latex beads. Exclusion of p67(phox) was associated with a defect of both NOX2 assembly and production of reactive oxygen species (ROS) in response to wild type Mtb. In contrast, Mtb Ndk-AS, which lost the capacity to disrupt Rac1-p67(phox) interaction, induced a strong ROS production. Given the established link between NOX2 activation and apoptosis, the proportion of Annexin V positive cells and levels of intracellular active caspase 3 were significantly higher in cells infected with Mtb Ndk-AS compared to wild type Mtb. Thus, knock down of Ndk converted Mtb into a pro-apoptotic mutant strain that has a phenotype of increased susceptibility to intracellular killing and reduced virulence in vivo. Taken together, our in vitro and in vivo data revealed that Ndk contributes significantly to Mtb virulence via attenuation of NADPH oxidase-mediated host innate immunity.
Assuntos
Proteínas de Bactérias/metabolismo; Imunidade Inata; Macrófagos/imunologia; Mycobacterium tuberculosis/imunologia; Neuropeptídeos/metabolismo; Núcleosídeo-Difosfato Quinase/metabolismo; Tuberculose Pulmonar/imunologia; Proteínas rac1 de Ligação ao GTP/metabolismo; Animais; Proteínas de Bactérias/antagonistas & inibidores; Proteínas de Bactérias/genética; Células da Medula Óssea/citologia; Células da Medula Óssea/enzimologia; Células da Medula Óssea/imunologia; Células da Medula Óssea/microbiologia; Linhagem Celular Transformada; Células Cultivadas; Feminino; Humanos; Macrófagos/enzimologia; Macrófagos/microbiologia; Macrófagos/ultraestrutura; Camundongos; Camundongos Endogâmicos C57BL; Camundongos SCID; Proteínas Mutantes/antagonistas & inibidores; Proteínas Mutantes/genética; Proteínas Mutantes/metabolismo; Mycobacterium tuberculosis/enzimologia; Mycobacterium tuberculosis/patogenicidade; Mycobacterium tuberculosis/fisiologia; NADPH Oxidases/antagonistas & inibidores; NADPH Oxidases/metabolismo; Neuropeptídeos/antagonistas & inibidores; Neuropeptídeos/genética; Núcleosídeo-Difosfato Quinase/antagonistas & inibidores; Núcleosídeo-Difosfato Quinase/genética; Oligorribonucleotídeos Antissenso; Fagossomos/enzimologia; Fagossomos/ultraestrutura; Proteínas Recombinantes/química; Proteínas Recombinantes/metabolismo; Tuberculose Pulmonar/enzimologia; Tuberculose Pulmonar/microbiologia; Tuberculose Pulmonar/patologia; Virulência; Proteínas rac1 de Ligação ao GTP/antagonistas & inibidores; Proteínas rac1 de Ligação ao GTP/genética

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Proteínas de Bactérias / Tuberculose Pulmonar / Neuropeptídeos / Núcleosídeo-Difosfato Quinase / Proteínas rac1 de Ligação ao GTP / Imunidade Inata / Macrófagos / Mycobacterium tuberculosis Tipo de estudo: Prognostic_studies Idioma: En Ano de publicação: 2013 Tipo de documento: Article

Texto completo: 1 Base de dados: MEDLINE Assunto principal: Proteínas de Bactérias / Tuberculose Pulmonar / Neuropeptídeos / Núcleosídeo-Difosfato Quinase / Proteínas rac1 de Ligação ao GTP / Imunidade Inata / Macrófagos / Mycobacterium tuberculosis Tipo de estudo: Prognostic_studies Idioma: En Ano de publicação: 2013 Tipo de documento: Article