TY - JOUR
T1 - Host resistance to intracellular infection
T2 - Mutation of natural resistance-associated macrophage protein 1 (Nramp1) impairs phagosomal acidification
AU - Hackam, David J.
AU - Rotstein, Ori D.
AU - Zhang, Wei Jian
AU - Gruenheid, Samantha
AU - Gros, Philippe
AU - Grinstein, Sergio
PY - 1998/7/20
Y1 - 1998/7/20
N2 - The mechanisms underlying the survival of intracellular parasites such as mycobacteria in host macrophages remain poorly understood. In mice, mutations at the Nramp1 gene (for natural resistance-associated macrophage protein), cause susceptibility to mycobacterial infections. Nramp1 encodes an integral membrane protein that is recruited to the phagosome membrane in infected macrophages. In this study, we used micro fluorescence ratio imaging of macrophages from wild-type and Nramp1 mutant mice to analyze the effect of loss of Nramp1 function on the properties of phagosomes containing inert particles or live mycobacteria. The pH of phagosomes containing live Mycobacterium bovis was significantly more acidic in Nramp1-expressing macrophages than in mutant cells (pH 5.5 ± 0.06 versus pH 6.6 ± 0.05, respectively; P <0.005). The enhanced acidification could not be accounted for by differences in proton consumption during dismutation of superoxide, phagosomal buffering power, counterion conductance, or in the rate of proton 'leak', as these were found to be comparable in wild-type and Nramp1- deficient macrophages. Rather, after ingestion of live mycobacteria, Nramp1- expressing cells exhibited increased concanamycin-sensitive H+ pumping across the phagosomal membrane. This was associated with an enhanced ability of phagosomes to fuse with vacuolar-type ATPase-containing late endosomes and/or lysosomes. This effect was restricted to live M. bovis and was not seen in phagosomes containing dead M. bovis or latex beads. These data support the notion that Nramp1 affects intracellular mycobacterial replication by modulating phagosomal pH, suggesting that Nramp1 plays a central role in this process.
AB - The mechanisms underlying the survival of intracellular parasites such as mycobacteria in host macrophages remain poorly understood. In mice, mutations at the Nramp1 gene (for natural resistance-associated macrophage protein), cause susceptibility to mycobacterial infections. Nramp1 encodes an integral membrane protein that is recruited to the phagosome membrane in infected macrophages. In this study, we used micro fluorescence ratio imaging of macrophages from wild-type and Nramp1 mutant mice to analyze the effect of loss of Nramp1 function on the properties of phagosomes containing inert particles or live mycobacteria. The pH of phagosomes containing live Mycobacterium bovis was significantly more acidic in Nramp1-expressing macrophages than in mutant cells (pH 5.5 ± 0.06 versus pH 6.6 ± 0.05, respectively; P <0.005). The enhanced acidification could not be accounted for by differences in proton consumption during dismutation of superoxide, phagosomal buffering power, counterion conductance, or in the rate of proton 'leak', as these were found to be comparable in wild-type and Nramp1- deficient macrophages. Rather, after ingestion of live mycobacteria, Nramp1- expressing cells exhibited increased concanamycin-sensitive H+ pumping across the phagosomal membrane. This was associated with an enhanced ability of phagosomes to fuse with vacuolar-type ATPase-containing late endosomes and/or lysosomes. This effect was restricted to live M. bovis and was not seen in phagosomes containing dead M. bovis or latex beads. These data support the notion that Nramp1 affects intracellular mycobacterial replication by modulating phagosomal pH, suggesting that Nramp1 plays a central role in this process.
KW - Macrophage
KW - Mycobacterium tuberculosis
KW - Phagocytosis
KW - Phagosome
KW - Proton pump
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U2 - 10.1084/jem.188.2.351
DO - 10.1084/jem.188.2.351
M3 - Article
C2 - 9670047
AN - SCOPUS:0031850526
SN - 0022-1007
VL - 188
SP - 351
EP - 364
JO - Journal of Experimental Medicine
JF - Journal of Experimental Medicine
IS - 2
ER -