Brefeldin A redistributes resident and itinerant Golgi proteins to the endoplasmic reticulum.
Brefeldin A redistributes resident and itinerant Golgi proteins to the endoplasmic reticulum.
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DOI：
10.1083/jcb.109.1.61
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发表时间：
1989-07
期刊：
影响因子：
--
通讯作者：
Yewdell JW
中科院分区：
文献类型：
--
作者：
Doms RW;Russ G;Yewdell JW
布雷菲德菌素A（Brefeldin A，BFA）可阻断蛋白质从内质网的转运，并导致高尔基复合体解体。我们研究了BFA对水泡性口炎病毒G蛋白（一种模型整合膜蛋白）的运输和加工的影响。6微克/毫升的BFA可逆地阻断G蛋白向细胞表面的递送。脉冲标记实验表明，在BFA的存在下，G蛋白变得完全耐内切糖苷酶H消化。添加唾液酸，一个trans-Golgi事件，没有观察到。尽管处理的顺式和内侧高尔基体酶，G蛋白被本地化的间接免疫荧光的ER的网状分布特征。通过用代谢抑制剂羰基氰化物间氯苯腙或通过使用温度敏感突变体ts 045（其在40 ° C下限于ER）阻止G蛋白从ER转运，我们表明G蛋白的加工发生在ER中，并且不是由于新合成的高尔基体酶的保留。相反，重新分配预先存在的顺式和内侧高尔基体酶的ER发生后2.5分钟，加入BFA，并完成了10-15分钟。高尔基体酶的ER交付是能量依赖性的，只发生在温度大于或等于20摄氏度。BFA还诱导G蛋白从内侧高尔基体向内质网的逆行转运。去除BFA后10分钟，高尔基体酶从ER中完全回收。这些研究结果表明，BFA诱导逆行运输居民和巡回高尔基体蛋白质的ER在一个完全可逆的方式。
Brefeldin A (BFA) has been reported to block protein transport from the ER and cause disassembly of the Golgi complex. We have examined the effects of BFA on the transport and processing of the vesicular stomatitis virus G protein, a model integral membrane protein. Delivery of G protein to the cell surface was reversibly blocked by 6 micrograms/ml BFA. Pulse-label experiments revealed that in the presence of BFA, G protein became completely resistant to endoglycosidase H digestion. Addition of sialic acid, a trans-Golgi event, was not observed. Despite processing by cis- and medial Golgi enzymes, G protein was localized by indirect immunofluorescence to a reticular distribution characteristic of the ER. By preventing transport of G protein from the ER with the metabolic inhibitor carbonyl cyanide m-chlorophenylhydrazone or by use of the temperature- sensitive mutant ts045, which is restricted to the ER at 40 degrees C, we showed that processing of G protein occurred in the ER and was not due to retention of newly synthesized Golgi enzymes. Rather, redistribution of preexisting cis and medial Golgi enzymes to the ER occurred as soon as 2.5 min after addition of BFA, and was complete by 10-15 min. Delivery of Golgi enzymes to the ER was energy dependent and occurred only at temperatures greater than or equal to 20 degrees C. BFA also induced retrograde transport of G protein from the medial Golgi to the ER. Golgi enzymes were completely recovered from the ER 10 min after removal of BFA. These findings demonstrate that BFA induces retrograde transport of both resident and itinerant Golgi proteins to the ER in a fully reversible manner.