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Targeting of the GTPase Irgm1 to the phagosomal membrane via PtdIns(3,4)P(2) and PtdIns(3,4,5)P(3) promotes immunity to mycobacteria.

ABSTRACT: Vertebrate immunity to infection enlists a newly identified family of 47-kilodalton immunity-related GTPases (IRGs). One IRG in particular, Irgm1, is essential for macrophage host defense against phagosomal pathogens, including Mycobacterium tuberculosis (Mtb). Here we show that Irgm1 targets the mycobacterial phagosome through lipid-mediated interactions with phosphatidylinositol-3,4-bisphosphate (PtdIns(3,4)P(2)) and PtdIns(3,4,5)P(3). An isolated Irgm1 amphipathic helix conferred lipid binding in vitro and in vivo. Substitutions in this region blocked phagosome recruitment and failed to complement the antimicrobial defect in Irgm1(-/-) macrophages. Removal of PtdIns(3,4,5)P(3) or inhibition of class I phosphatidylinositol-3-OH kinase (PI(3)K) mimicked this effect in wild-type cells. Cooperation between Irgm1 and PI(3)K further facilitated the engagement of Irgm1 with its fusogenic effectors at the site of infection, thereby ensuring pathogen-directed responses during innate immunity.

SUBMITTER: Tiwari S

PROVIDER: S-EPMC2715447 | biostudies-literature | 2009 Aug

REPOSITORIES: biostudies-literature

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Targeting of the GTPase Irgm1 to the phagosomal membrane via PtdIns(3,4)P(2) and PtdIns(3,4,5)P(3) promotes immunity to mycobacteria.

Tiwari Sangeeta S Choi Han-Pil HP Matsuzawa Takeshi T Pypaert Marc M MacMicking John D JD

Nature immunology 20090801 8

Vertebrate immunity to infection enlists a newly identified family of 47-kilodalton immunity-related GTPases (IRGs). One IRG in particular, Irgm1, is essential for macrophage host defense against phagosomal pathogens, including Mycobacterium tuberculosis (Mtb). Here we show that Irgm1 targets the mycobacterial phagosome through lipid-mediated interactions with phosphatidylinositol-3,4-bisphosphate (PtdIns(3,4)P(2)) and PtdIns(3,4,5)P(3). An isolated Irgm1 amphipathic helix conferred lipid bindin ...[more]

PMID: 19620982

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Project description:Many cytosolic proteins are recruited to the plasma membrane (PM) during cell signaling and other cellular processes. Recent reports have indicated that phosphatidylserine (PS), phosphatidylinositol 4,5-bisphosphate (PtdIns(4,5)P(2)), and phosphatidylinositol 3,4,5-trisphosphate (PtdIns(3,4,5)P(3)) that are present in the PM play important roles for their specific PM recruitment. To systematically analyze how these lipids mediate PM targeting of cellular proteins, we performed biophysical, computational, and cell studies of the Ca(2+)-dependent C2 domain of protein kinase Calpha (PKCalpha) that is known to bind PS and phosphoinositides. In vitro membrane binding measurements by surface plasmon resonance analysis show that PKCalpha-C2 nonspecifically binds phosphoinositides, including PtdIns(4,5)P(2) and PtdIns(3,4,5)P(3), but that PS and Ca(2+) binding is prerequisite for productive phosphoinositide binding. PtdIns(4,5)P(2) or PtdIns(3,4,5)P(3) augments the Ca(2+)- and PS-dependent membrane binding of PKCalpha-C2 by slowing its membrane dissociation. Molecular dynamics simulations also support that Ca(2+)-dependent PS binding is essential for membrane interactions of PKCalpha-C2. PtdIns(4,5)P(2) alone cannot drive the membrane attachment of the domain but further stabilizes the Ca(2+)- and PS-dependent membrane binding. When the fluorescence protein-tagged PKCalpha-C2 was expressed in NIH-3T3 cells, mutations of phosphoinositide-binding residues or depletion of PtdIns(4,5)P(2) and/or PtdIns(3,4,5)P(3) from PM did not significantly affect the PM association of the domain but accelerated its dissociation from PM. Also, local synthesis of PtdIns(4,5)P(2) or PtdIns(3,4,5)P(3) at the PM slowed membrane dissociation of PKCalpha-C2. Collectively, these studies show that PtdIns(4,5)P(2) and PtdIns(3,4,5)P(3) augment the Ca(2+)- and PS-dependent membrane binding of PKCalpha-C2 by elongating the membrane residence of the domain but cannot drive the PM recruitment of PKCalpha-C2. These studies also suggest that effective PM recruitment of many cellular proteins may require synergistic actions of PS and phosphoinositides.

Dataset Information

Targeting of the GTPase Irgm1 to the phagosomal membrane via PtdIns(3,4)P(2) and PtdIns(3,4,5)P(3) promotes immunity to mycobacteria.

Publications

Targeting of the GTPase Irgm1 to the phagosomal membrane via PtdIns(3,4)P(2) and PtdIns(3,4,5)P(3) promotes immunity to mycobacteria.

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