Proton-activated chloride channel PAC regulates endosomal acidification and transferrin receptor-mediated endocytosis

During vesicular acidification, chloride (Cl^-), as the counterion, provides the electrical shunt for proton pumping by the vacuolar H⁺ ATPase. Intracellular CLC transporters mediate Cl^- influx to the endolysosomes through their 2Cl^-/H⁺ exchange activity. However, whole-endolysosomal patch-clamp recording also revealed a mysterious conductance releasing Cl^- from the lumen. It remains unknown whether CLCs or other Cl^- channels are responsible for this activity. Here, we show that the newly identified proton-activated Cl^- (PAC) channel traffics from the plasma membrane to endosomes via the classical YxxL motif. PAC deletion abolishes the endosomal Cl^- conductance, raises luminal Cl^- level, lowers luminal pH, and increases transferrin receptor-mediated endocytosis. PAC overexpression generates a large endosomal Cl^- current with properties similar to those of endogenous conductance, hypo-acidifies endosomal pH, and reduces transferrin uptake. We propose that the endosomal Cl^- PAC channel functions as a low pH sensor and prevents hyper-acidification by releasing Cl^- from the lumen.