Electrogenic nature of lysosomal proton pump as revealed with a cyanine dye.
Studies were carried out on the electrogenicity of the lysosomal proton pump using dipropylthiadicarbocyanine iodide (diS-C3-(5] as a membrane potential probe. Pure lysosome preparations (tritosomes) quenched the fluorescence of diS-C3-(5). The quenching correlated well with the potassium ion diffusion potential (inside negative) generated by K+ with or without valinomycin. The quenching caused by lysosomes was reversed by lipophilic cations, tetraphenylarsonium (TPA) or triphenylmethylphosphonium (TPMP). Mg-ATP also reversed the quenching, which was inhibited by a protonophore, 3,5-di-tert-butyl-4-hydroxybenzylidene-malononitrile (SF-6847). The properties of the ATP-induced recovery of the quenching were exactly the same as those of ATP-induced acidification, as measured with fluorescein isothiocyanate-dextran (FD) (Ohkuma, S., et al. (1982) Proc. Natl. Acad. Sci. U.S. 79, 2758-2762) and acridine orange (Moriyama, Y., et al. (1982) J. Biochem. 92, 1333-1336), except replacement of the anion by an impermeable one enhanced ATP-induced recovery of quenching, but reduced ATP-induced acidification. Amines which dissipate delta pH across the lysosomal membrane also enhanced the Mg-ATP-induced fluorescence recovery. These results suggest that isolated lysosomes exhibit an inside negative membrane potential, especially in low K+ medium, mostly due to the K+-diffusion potential, and that the Mg-ATP-driven proton pump causes membrane depolarization (in the direction of inside positive). These possibilities were supported by results on the uptake of the radioactive membrane-permeant ions [3H]TPMP and [14C]SCN. The present results provide evidence for the electrogenic nature of the lysosomal proton pump.
D000255:Adenosine Triphosphate / Q000494:pharmacology; D000818:Animals; D052160:Benzothiazoles; D001693:Biological Transport, Active / Q000187:drug effects; D002232:Carbocyanines; D005456:Fluorescent Dyes; D006863:Hydrogen-Ion Concentration; D066298:In Vitro Techniques; D007700:Kinetics; D008099:Liver / Q000378:metabolism*; D008247:Lysosomes / Q000378:metabolism*; D008297:Male; D011188:Potassium / Q000494:pharmacology; D011804:Quinolines; D051381:Rats; D011919:Rats, Inbred Strains
D052160:Benzothiazoles; D002232:Carbocyanines; D005456:Fluorescent Dyes; D011804:Quinolines; C012944:3,3'-dipropyl-2,2'-thiadicarbocyanine; D000255:Adenosine Triphosphate; D011188:Potassium