Plasma membrane-associated actin in bright yellow 2 tobacco cells - Evidence for interaction with microtubules
Plasma membrane-associated actin in bright yellow 2 tobacco cells - Evidence for interaction with microtubules
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DOI：
10.1104/pp.118.3.917
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发表时间：
1998-11-01
期刊：
影响因子：
7.4
通讯作者：
Shibaoka, H
中科院分区：
文献类型：
--
作者：
Collings, DA;Asada, T;Shibaoka, H
当附着在底物上的原生质体被裂解后留下一小块质膜时，就会形成质膜幽灵。我们已经确定了几个因素，包括使用轻度酸性肌动蛋白稳定缓冲液和在固定液中包含戊二醛，这些因素允许免疫荧光可视化保留在烟草（Nicotiana tabacum L.）悬浮培养细胞（Blight Yellow 2系）的膜上的广泛皮层肌动蛋白阵列。在这些条件下也保留了正常的微管阵列。膜相关肌动蛋白是随机的；它与微管仅表现出有限的协同作用，微管在细胞壁消化和鬼影形成之前在整个细胞中解聚对肌动蛋白保留几乎没有影响。肌动蛋白和微管对裂解缓冲液的pH、K+和Ca2+浓度也表现出不同的敏感性。然而，有强有力的证据表明肌动蛋白和微管在质膜或质膜附近相互作用，因为由紫杉醇预处理的细胞制备的鬼细胞和原生质体都有平行排列的微管，并且与微管对齐的肌动蛋白数量增加。这些实验表明，皮层肌动蛋白阵列的组织可能取决于微管的定位和组织。
Plasma membrane ghosts form when plant protoplasts attached to a substrate are lysed to leave a small patch of plasma membrane. We have identified several factors, including the use of a mildly acidic actin stabilization buffer and the inclusion of glutaraldehyde in the fixative, that allow immunofluorescent visualization of extensive cortical actin arrays retained on membrane ghosts made from tobacco (Nicotiana tabacum L.) suspension-cultured cells (line Blight Yellow 2). Normal microtubule arrays were also retained using these conditions. Membrane-associated actin is random; it exhibits only limited coalignment with the microtubules, and microtubule depolymerization in whole cells before wall digestion and ghost formation has little effect on actin retention. Actin and microtubules also exhibit different sensitivities to the pH and K+ and Ca2+ concentrations of the lysis buffer. There is, however, strong evidence for interactions between actin and the microtubules at or near the plasma membrane, because both ghosts and protoplasts prepared from taxol-pretreated cells have microtubules arranged in parallel arrays and an increased amount of actin coaligned with the microtubules. These experiments suggest that the organization of the cortical actin arrays may be dependent on the localization and organization of the microtubules.