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Updated: Jun 14, 2026
Measuring Phagosome pH by Ratiometric Fluorescence Microscopy
Published on: December 7, 2015
Francisella acid phosphatases inactivate the NADPH oxidase in human phagocytes
Nrusingh P Mohapatra1, Shilpa Soni, Murugesan V S Rajaram
1Department of Molecular Virology, Immunology and Medical Genetics, Center for Microbial Interface Biology, The Ohio State University, Columbus, OH 43210, USA.
Abstract:
Francisella tularensis contains four putative acid phosphatases that are conserved in Francisella novicida. An F. novicida quadruple mutant (AcpA, AcpB, AcpC, and Hap [DeltaABCH]) is unable to escape the phagosome or survive in macrophages and is attenuated in the mouse model. We explored whether reduced survival of the DeltaABCH mutant within phagocytes is related to the oxidative response by human neutrophils and macrophages. F. novicida and F. tularensis subspecies failed to stimulate reactive oxygen species production in the phagocytes, whereas the F. novicida DeltaABCH strain stimulated a significant level of reactive oxygen species. The DeltaABCH mutant, but not the wild-type strain, strongly colocalized with p47(phox) and replicated in phagocytes only in the presence of an NADPH oxidase inhibitor or within macrophages isolated from p47(phox) knockout mice. Finally, purified AcpA strongly dephosphorylated p47(phox) and p40(phox), but not p67(phox), in vitro. Thus, Francisella acid phosphatases play a major role in intramacrophage survival and virulence by regulating the generation of the oxidative burst in human phagocytes.
Insights
Francisella acid phosphatases are crucial for bacterial survival within host cells. These enzymes help Francisella bacteria evade the oxidative burst, a key immune response, enabling them to replicate inside macrophages.
Area of Science:
- Microbiology
- Immunology
- Bacterial Pathogenesis
Background:
- Francisella tularensis is an intracellular bacterium that causes tularemia.
- Acid phosphatases are enzymes involved in various cellular processes.
- Francisella species possess four putative acid phosphatases conserved between F. novicida and F. tularensis.
Purpose of the Study:
- To investigate the role of Francisella acid phosphatases in bacterial survival and virulence within phagocytic cells.
- To determine if acid phosphatases influence the host's oxidative response.
Main Methods:
- Construction and characterization of a quadruple acid phosphatase mutant (DeltaABCH) of F. novicida.
- Assessment of reactive oxygen species (ROS) production in human neutrophils and macrophages upon infection with wild-type and mutant F. novicida.
- Analysis of bacterial colocalization with p47(phox) and replication in phagocytes under different conditions (NADPH oxidase inhibition, p47(phox) knockout macrophages).
- In vitro dephosphorylation assays using purified AcpA enzyme.
Main Results:
- The F. novicida DeltaABCH mutant failed to escape the phagosome, showed reduced survival in macrophages, and was attenuated in mice.
- Wild-type F. novicida did not induce ROS production, while the DeltaABCH mutant stimulated a significant ROS response.
- The DeltaABCH mutant colocalized with p47(phox) and replicated in phagocytes only when the oxidative burst was inhibited or absent.
- Purified AcpA dephosphorylated p47(phox) and p40(phox) in vitro.
Conclusions:
- Francisella acid phosphatases are essential virulence factors that promote intramacrophage survival.
- These enzymes counteract the host's oxidative burst by dephosphorylating key components of the NADPH oxidase complex, such as p47(phox).
- Targeting Francisella acid phosphatases could be a potential therapeutic strategy against tularemia.
