Bafilomycin A1, a specific inhibitor of vacuolar-type H(+)-ATPase, inhibits acidification and protein degradation in lysosomes of cultured cells.
Bafilomycin A1, a specific inhibitor of vacuolar-type H(+)-ATPase, inhibits acidification and protein degradation in lysosomes of cultured cells.
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DOI：
10.1016/s0021-9258(19)47429-2
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发表时间：
1991-09
期刊：
影响因子：
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通讯作者：
T. Yoshimori;A. Yamamoto;Yoshinori MoriyamaS;Masamitsu FutaiS;Yutaka TashiroQ
中科院分区：
文献类型：
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作者：
T. Yoshimori;A. Yamamoto;Yoshinori MoriyamaS;Masamitsu FutaiS;Yutaka TashiroQ
众所周知，巴菲霉素A1在体外是空泡型H(+)- atp酶的强抑制剂，而其他类型的atp酶，如F1， f0 - atp酶，不受这种抗生素的影响(Bowman, e.m., Siebers, a ., and Altendorf， K. (1988) Proc. Natl。学会科学。（美国法典85,7972-7976）。检测了该抑制剂对活培养细胞溶酶体的影响。0.1-1 μ m巴菲霉素A1处理BNL cl - 2和A431细胞，可完全抑制吖啶橙孵育显示的溶酶体酸化。这种效果是通过清洗细胞显现出来的。使用3-（2,4-二硝基苯胺）-3′-氨基- n -甲基二丙胺和异硫氰酸-葡聚糖荧光素进行的两项研究表明，在1微米的巴菲霉素A1存在下，A431细胞的溶体内pH从约5.1-5.5增加到约6.3。pH值在50 min左右逐渐升高。在1 μ m巴菲霉素A1存在的情况下，在4℃下与细胞表面结合的125i标记的表皮生长因子（EGF）在37℃下正常内化到细胞中，但根本不降解，与没有药物的对照细胞中125I-EGF的快速降解形成鲜明对比。免疫金电镜显示，无论是否添加巴菲霉素A1， EGF都被转运到溶酶体中。这些结果表明，液泡型H(+)- atp酶在体内溶酶体的酸化和蛋白质降解中起关键作用。
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 microM bafilomycin A1. The effect was revealed by washing the cells. Both studies using 3-(2,4-dinitroanilino)-3'-amino-N-methyldipropylamine and fluorescein isothiocyanate-dextran showed that the intralysomal pH of A431 cells increased from about 5.1-5.5 to about 6.3 in the presence of 1 microM bafilomycin A1. The pH increased gradually in about 50 min. In the presence of 1 microM bafilomycin A1, 125I-labeled epidermal growth factor (EGF) bound to the cell surface at 4 degrees C was internalized normally into the cells at 37 degrees 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(+)-ATPase plays a pivotal role in acidification and protein degradation in the lysosomes in vivo.