ENDOLYSOSOMAL TRPML3 / BK COMPLEX IN AUTOPHAGY INDUCTION AND PATHOGEN DEFENSE

Abstract

The transient receptor potential mucolipin 3 (TRPML3), encoded by the MCOLN3 gene, functions as a calcium (Ca²⁺) release channel localized to endolysosomal membranes. This channel is activated by phosphatidylinositol 3,5-bisphosphate (PI3,5P2), with its activity further enhanced by elevated luminal pH or substitution of luminal sodium (Na⁺) with potassium (K⁺). In this study, we identify a positive feedback loop between TRPML3 and the big-conductance Ca²⁺-activated potassium channel (BK). Ca²⁺ release through TRPML3 activates BK channels, which subsequently enhance TRPML3-mediated Ca²⁺ release, likely by providing a counter flux of K⁺ and alleviating the inhibitory effect of luminal Na⁺. We further demonstrate that TRPML3/BK and the mammalian target of rapamycin (mTOR) form another positive feedback loop. mTOR inhibition during nutrient starvation activates TRPML3/BK channels, which further suppress mTOR activity, thereby increasing autophagy induction. Mechanistically, this regulatory interplay is mediated by phosphatidylinositol 3-phosphate (PI3P), an endogenous TRPML3 activator that is enriched in phagophores and becomes upregulated as mTOR activity decreases. Importantly, bacterial infection triggers TRPML3 activation in a BK-dependent manner, with both TRPML3 and BK playing crucial roles in the suppression of mTOR and the promotion of autophagy in response to infection. Inhibition of TRPML3 or BK enhances intracellular bacterial survival, while upregulation of either protein facilitates bacterial clearance. Given that TRPML3/BK is inhibited at low luminal pH but activated by high luminal pH and PI3P in phagophores, we suggest that the TRPML3/BK and mTOR feedback loop via PI3P is critical for efficient autophagy induction during nutrient deprivation or bacterial infection. These findings reveal a pivotal role for TRPML3-BK coupling in maintaining cellular homeostasis and mediating intracellular bacterial clearance by regulating mTOR signaling pathways.