Cholinergic agonist PNU282987 induces pro-inflammatory gene expression via the MCOLN1-TFEB pathway in murine macrophages.

Abstract

Transcription factors TFEB and TFE3 are crucial for regulating autophagy, lysosomal biogenesis, and lipid metabolism, and have significant roles in macrophage function and innate immunity. The alpha7 nicotinic acetylcholine receptor (7nAChR), a ligand-gated Ca2+ channel known for its therapeutic potential in neurological and inflammatory disorders, has been implicated in modulating immune responses by modulating macrophage function. Stimulation of 7nAChR with chemical agonists has been claimed to activate TFEB in pancreatic acinar cells and neurons. However, the impact of 7nAChR activation on TFEB and TFE3 in macrophages remained unknown, posing an important question due to the potential implications for inflammation regulation. This study investigates the effects of acute 7nAChR activation on TFEB-mediated responses in murine macrophages using the specific agonist PNU-282987. We demonstrate that 7nAChR stimulation triggers TFEB nuclear translocation and lysosomal expansion. Surprisingly, PNU-282987 induces a broad pro-inflammatory gene signature without concomitant cytokine secretion, suggesting an uncoupling of gene expression from cytokine release. Mechanistically, TFEB activation requires the lysosomal Ca2+ exporter MCOLN1 and the Ca2+-dependent phosphatase PPP3/calcineurin. Additionally, PNU-282987 elevates reactive oxygen species (ROS) levels, and ROS are involved in TFEB activation by PNU-282987. Notably, even with 7nAChR deletion, compensatory ROS-mediated TFEB activation persists, suggesting the involvement of additional nicotinic receptors. Our findings reveal a novel 7nAChR-TFEB signaling axis in macrophages, offer new insights into the cholinergic regulation of immune responses, establish a baseline for comparison with disease states, and identify potential therapeutic targets for modulating inflammation.