Bafilomycin A1 is known as a strong inhibitor of the vacuolar type H+-ATPase in vitro, whereas other type ATPases, e.g. F1,F0-ATPase, are not affected by this antibiotic (Bowman, E. M., Siebers, A., and Altendorf, K. (1988) Proc. Natl. Acad. Sci. U. S. A. 85, 7972-7976). Effects of this inhibitor on lysosomes of living cultured cells were tested. The acidification of lysosomes revealed by the incubation with acridine orange was completely inhibited when BNL CL.2 and A431 cells were treated with 0.1-1 μM bafilomycin A1. The effect was reversed by washing the cells. Both studies using 3-(2,4-dinitroanilino)-3′-amino-N- methyldipropylamine and fluorescein isothiocyanate-dextran showed that the intralysosomal pH of A431 cells increased from about 5.1-5.5 to about 6.3 in the presence of 1 μM bafilomycin A1. The pH increased gradually in about 50 min. In the presence of 1 μM bafilomycin A1, 125I-labeled epidermal growth factor (EGF) bound to the cell surface at 4 °C was internalized normally into the cells at 37 °C but was not degraded at all, in marked contrast to the rapid degradation of 125I-EGF in the control cells without the drug. Immunogold electron microscopy showed that EGF was transported into lysosomes irrespective of the addition of bafilomycin A1. These results suggest that the vacuolar type H+-ATP-ase plays a pivotal role in acidification and protein degradation in the lysosomes in vivo.
|Number of pages||6|
|Journal||Journal of Biological Chemistry|
|Publication status||Published - Sep 15 1991|
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