title: The glyceraldehyde-3-phosphate dehydrogenase and the small GTPase Rab 2 are crucial for Brucella replication.
abstract: The intracellular pathogen Brucella abortus survives and replicates inside host cells within an endoplasmic reticulum (ER)-derived replicative organelle named the "Brucella-containing vacuole" (BCV). Here, we developed a subcellular fractionation method to isolate BCVs and characterize for the first time the protein composition of its replicative niche. After identification of BCV membrane proteins by 2 dimensional (2D) gel electrophoresis and mass spectrometry, we focused on two eukaryotic proteins: the glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the small GTPase Rab 2 recruited to the vacuolar membrane of Brucella. These proteins were previously described to localize on vesicular and tubular clusters (VTC) and to regulate the VTC membrane traffic between the endoplasmic reticulum (ER) and the Golgi. Inhibition of either GAPDH or Rab 2 expression by small interfering RNA strongly inhibited B. abortus replication. Consistent with this result, inhibition of other partners of GAPDH and Rab 2, such as COPI and PKC iota, reduced B. abortus replication. Furthermore, blockage of Rab 2 GTPase in a GDP-locked form also inhibited B. abortus replication. Bacteria did not fuse with the ER and instead remained in lysosomal-associated membrane vacuoles. These results reveal an essential role for GAPDH and the small GTPase Rab 2 in B. abortus virulence within host cells.
authors: Aix Marseille Université, Centre d'Immunologie de Marseille-Luminy, UMR6546, Marseille, France.
authors: Emilie
authors: E
authors: Fugier
authors: Suzana P
authors: SP
authors: Salcedo
authors: Chantal
authors: C
authors: de Chastellier
authors: Matthieu
authors: M
authors: Pophillat
authors: Alexandre
authors: A
authors: Muller
authors: Vilma
authors: V
authors: Arce-Gorvel
authors: Patrick
authors: P
authors: Fourquet
authors: Jean-Pierre
authors: JP
authors: Gorvel
mesh_terms: D000818:Animals; D002003:Brucella abortus / Q000166:cytology* / Q000254:growth & development; D002460:Cell Line; D002462:Cell Membrane / Q000737:chemistry / Q000382:microbiology; D002470:Cell Survival; D004721:Endoplasmic Reticulum / Q000382:microbiology; D024581:Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating) / Q000737:chemistry / Q000378:metabolism*; D054884:Host-Pathogen Interactions / Q000502:physiology; D007150:Immunohistochemistry; D008565:Membrane Proteins / Q000737:chemistry / Q000378:metabolism; D055571:Secretory Pathway / Q000502:physiology; D014617:Vacuoles / Q000737:chemistry / Q000201:enzymology / Q000382:microbiology; D020693:rab2 GTP-Binding Protein / Q000737:chemistry / Q000378:metabolism*
chemical_list: D008565:Membrane Proteins; D024581:Glyceraldehyde-3-Phosphate Dehydrogenase (Phosphorylating); D020693:rab2 GTP-Binding Protein