Protein phosphatase 1 regulates the cytoplasmic dynein-driven formation of endoplasmic reticulum networks in vitro.
Protein phosphatase 1 regulates the cytoplasmic dynein-driven formation of endoplasmic reticulum networks in vitro.
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DOI：
10.1083/jcb.128.5.879
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发表时间：
1995-03
期刊：
影响因子：
--
通讯作者：
Allan V
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文献类型：
--
作者：
Allan V
间期非洲爪蟾卵提取物在体外形成广泛的管状膜网络。这些网络在这里通过内质网驻留蛋白的存在（如免疫荧光所示）以及单个核糖体和多核糖体的存在（如电子显微镜所示）被鉴定为内质网。使用磷酸酶抑制剂冈田酸测试磷酸化对间期内质网运动的影响。与对照提取物相比，冈田酸处理导致 ER 小管移动数量和形成 ER 网络的范围增加了 27 倍。这种激活被广泛特异性激酶抑制剂 6-二甲氨基嘌呤阻断。然而，冈田酸对内质网小管运动的方向没有影响，内质网小管运动发生在微管的负端，并且对低浓度的钒酸盐敏感。磷酸酶的抑制也对 ER 小管延伸的速度或持续时间没有影响，并且不会刺激可溶性细胞质动力蛋白的活性。内质网运动对冈田酸的敏感性与蛋白磷酸酶 1 的敏感性密切相关。尽管通过抑制蛋白磷酸酶 1 (PP1) 大大增加了内质网运动量，但与膜相关的细胞质动力蛋白的量并未改变。这些数据支持这样一个模型：磷酸化通过控制与内质网膜结合的细胞质动力蛋白的活性来调节内质网运动。
Interphase Xenopus egg extracts form extensive tubular membrane networks in vitro. These networks are identified here as endoplasmic reticulum by the presence of ER resident proteins, as shown by immunofluorescence, and by the presence of single ribosomes and polysomes, as shown by electron microscopy. The effect of phosphorylation on ER movement in interphase was tested using the phosphatase inhibitor, okadaic acid. Okadaic acid treatment resulted in an increase of up to 27-fold in the number of ER tubules moving and in the extent of ER networks formed compared to control extracts. This activation was blocked by the broad-specificity kinase inhibitor 6- dimethylaminopurine. Okadaic acid had no effect, however, on the direction of ER tubule movement, which occurred towards the minus end of microtubules, and was sensitive to low concentrations of vanadate. Inhibition of phosphatases also had no effect on the speed or duration of ER tubule extensions, and did not stimulate the activity of soluble cytoplasmic dynein. The sensitivity of ER movement to okadaic acid closely matched that of protein phosphatase 1. Although the amount of ER motility was greatly increased by inhibiting protein phosphatase 1 (PP1), the amount of cytoplasmic dynein associated with the membrane was not altered. The data support a model in which phosphorylation regulates ER movement by controlling the activity of cytoplasmic dynein bound to the ER membrane.