The Journal of Immunology
Autophagy is an evolutionarily conserved cytoplasmic process regulated by the energy rheostats mammalian target of rapamycin and AMP kinase (AMPK) that recycles damaged or unused proteins and organelles. It has been described as an important effector arm of immune cells. We have shown that the cytoplasmically oriented calcium/calmodulin-dependent protein kinase (CaMK)I regulates the inflammatory phenotype of the macrophage (M). In this study, we hypothesize that CaMKI mediates M autophagy. LPS induced autophagy in RAW 264.7 cells and murine peritoneal M that was attenuated with biochemical CaMK inhibition or CaMKI small interfering RNA (siRNA). Inhibition of CaMKI reduced LPS-induced p-Thr(172)AMPK and target of rapamycin complex-1 activity, and expression of a constitutively active CaMKI but not a kinase-deficient mutant induced p-Thr(172)AMPK and autophagy that was attenuated by the AMPK inhibitor compound C. Coimmunoprecipitation and in vitro kinase assays demonstrated that CaMKI activates AMPK, thereby inducing ATG7, which also localizes to this CaMKI/AMPK complex. During LPS-induced lung inflammation, C57BL/6 mice receiving CaMKI(siRNA) displayed reduced lung and bronchoalveolar immune cell autophagy that correlated with reduced neutrophil recruitment, myeloperoxidase activity, and air space cytokine concentration. Independently inhibiting autophagy, using siRNA targeting the PI3K VPS34, yielded similar reductions in lung autophagy and neutrophil recruitment. Thus, a novel CaMKI/AMPK pathway is rapidly activated in M exposed to LPS and regulates an early autophagic response, independent of target of rapamycin complex-1 inhibition. These mechanisms appear to be operant in vivo in orchestrating LPS-induced lung neutrophil recruitment and inflammation.