A NOD2-Encoded Toggle Switch Resolves the Host-Microbe Battle Over Cyclic AMP Control

Anandachar, M. S.; Chen, D.; Perkins, K. C.; Katkar, G. D.; Roy, S.; Espinoza, C. R.; Mullick, M.; Sinha, S.; Nakayama, M.; Salem, J.; Estanol, M.; Bhattacharjee, R.; Zablan, K.; Hsu, S.-T.; Williams, S.; Sun, A.; Tindle, C.; Yang, J.; Kufareva, I.; Ghosh, P.

doi:10.64898/2026.03.29.715116

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bioRxiv · 10.64898/2026.03.29.715116

A NOD2-Encoded Toggle Switch Resolves the Host-Microbe Battle Over Cyclic AMP Control

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Abstract

Pathogens hijack macrophages by triggering pathological cAMP surges that block phagolysosomal killing--a defect mirrored in phagocytes from refractory colitis. We identify a host-encoded, pathogen-specific surge-protector comprised of a three-protein toggle: The innate sensor NOD2 binds and masks an evolutionarily conserved motif in GIV that activates trimeric-GTPase Gi, enforcing a biphasic surge-to-plunge cAMP-program: early, NOD2*GIV assembly permits a brief, tolerogenic cAMP rise, whereas subsequent GIV*Gi engagement collapses cAMP to drive phagolysosomal fusion and microbial clearance. Structural, biochemical, and ultrastructural analyses reveal how molecular toggling imposes precise spatial and temporal control. Pharmacogenomic perturbations pinpoint cAMP-PKA hyperactivation as the defining lesion in GIV-deficient macrophages. Functional studies in primary macrophages and human gut organoid co-cultures show that toggling the NOD2*GIV*Gi-axis is necessary and sufficient to convert tolerant macrophages into microbicidal machines that preserve mucosal barrier integrity. These findings uncover a druggable cAMP-control pathway with therapeutic promise in colitis. GRAPHIC ABSTRACT O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/715116v1_ufig1.gif" ALT="Figure 1"> View larger version (70K): org.highwire.dtl.DTLVardef@53cd40org.highwire.dtl.DTLVardef@1c83c82org.highwire.dtl.DTLVardef@fcbc58org.highwire.dtl.DTLVardef@b14f8e_HPS_FORMAT_FIGEXP M_FIG C_FIG eTOC BlurbPathogens hijack macrophages by inducing cAMP surges that help them evade clearance. Anandachar et al. identify a host "toggle switch" in which NOD2 and G proteins compete for GIV, driving a rapid and robust surge-to-plunge transition in cAMP. This temporal switch limits tolerogenic signaling, restores microbial clearance and barrier integrity, and unveils a targetable host pathway in infection and IBD. HighlightsO_LIPathogens exploit cAMP surges in macrophages to block phagolysosomal killing of microbes C_LIO_LIGIV acts as a molecular "toggle" linking NOD2 sensing to Gi-mediated cAMP control C_LIO_LIStructural and mutagenesis studies reveal mutually exclusive binding of NOD2 and Gi to GIV C_LIO_LIPharmacogenomic perturbations pinpoint PKA, not EPAC, as the critical downstream effector C_LIO_LIOrganoid co-cultures show NOD2-GIV-PKA crosstalk safeguards microbial clearance and gut barrier integrity C_LI

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Anandachar, M. S., Chen, D., Perkins, K. C., Katkar, G. D., Roy, S., Espinoza, C. R., Mullick, M., Sinha, S., Nakayama, M., Salem, J., Estanol, M., Bhattacharjee, R., Zablan, K., Hsu, S.-T., Williams, S., Sun, A., Tindle, C., Yang, J., Kufareva, I., Ghosh, P.. 2026-03-31. A NOD2-Encoded Toggle Switch Resolves the Host-Microbe Battle Over Cyclic AMP Control. https://doi.org/10.64898/2026.03.29.715116

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