Mycobacterium bovis BCG disrupts the interaction of Rab7 with RILP contributing to inhibition of phagosome maturation.
Phagosomes containing M. tuberculosis and M. bovis BCG interact normally with early endosomes but fail to fuse with late endosomes and lysosomes. Whereas many early events of mycobacterial phagosomes have been elucidated, the exact mechanism of the inhibition of fusion with lysosomes is still unclear. Several Rab GTPase proteins were shown to be involved in membrane fusion and vesicular transport. In particular, Rab7 associates with the phagosomal membrane and regulates the fusion between late endosomes and lysosomes. This function of Rab7 was shown to be mediated in epithelial cell models by the Rab7 effector RILP (Rab7-interacting lysosomal protein). However, the relevance of Rab7-RILP interaction to phagosome biogenesis in macrophage infected with mycobacteria is still unknown. In this study, cotransfection of RAW 264.7 cells with Rab7 and RILP revealed that Rab7-RILP interaction occurs in macrophages ingesting latex beads. Thereafter, this cell system model was used to demonstrate that infection with live but not killed M. bovis BCG inhibited RILP recruitment despite Rab7 acquisition by the phagosome. Further investigation using immobilized RILP to pull down active Rab7 (GTP-bound form) from macrophage lysates demonstrated that inactive Rab7 (GDP-bound form) predominates in cells infected with live BCG. In addition, cell-free system experiments demonstrated that BCG culture supernatant contains a factor that catalyzes the GTP/GDP switch on recombinant Rab7 molecules. Such a factor was shown to diffuse beyond BCG phagosomes and target other Rab7-positive compartments. These findings suggest that live mycobacteria express within the macrophage a Rab7 deactivating factor leading to abortion of RILP-mediated fusion with lysosomes.
Division of Infectious Diseases, Department of Medicine, University of British Columbia and Vancouver Costal Health Institute, Vancouver, British Columbia, Canada.
Jim
J
Sun
Ala-Eddine
AE
Deghmane
Hafid
H
Soualhine
Thomas
T
Hong
Cecilia
C
Bucci
Anna
A
Solodkin
Zakaria
Z
Hmama
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D004058:Diffusion; D008264:Macrophages / Q000166:cytology / Q000382:microbiology; D051379:Mice; D008810:Mice, Inbred C57BL; D050296:Microbial Viability; D009163:Mycobacterium bovis / Q000502:physiology*; D010588:Phagosomes / Q000378:metabolism* / Q000382:microbiology*; D011485:Protein Binding; D020691:rab GTP-Binding Proteins / Q000378:metabolism*; D000090802:rab7 GTP-Binding Proteins
D002352:Carrier Proteins; D000090802:rab7 GTP-Binding Proteins; C000716465:rab7 GTP-binding proteins, mouse; D020691:rab GTP-Binding Proteins