Phosphorylation of T-558 of moesin detected by site-specific antibodies in RAW264.7 macrophages
Phosphorylation of T-558 of moesin detected by site-specific antibodies in RAW264.7 macrophages
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DOI：
10.1006/bbrc.1996.1410
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发表时间：
1996-09-24
影响因子：
3.1
通讯作者：
Furthmayr, H
中科院分区：
文献类型：
--
作者：
Nakamura, F;Amieva, MR;Furthmayr, H
为了确定莫斯蛋白羧基末端结构域中的 (558)Thr 在血小板以外的细胞中是否被磷酸化，针对莫斯蛋白序列的化学磷酸化合成六肽 KYKpTLR 以及未磷酸化形式制备了兔磷酸化状态特异性抗体。亲和纯化的抗体群体对与 BSA 缀合的磷酸化或未修饰的肽具有特异性。通过蛋白质印迹分析在 RAW 巨噬细胞中检测到 Moesin 的位点特异性磷酸化，免疫荧光研究表明磷酸化的 moesin 位于丝状突起中。用磷酸酶抑制剂 calyculin A 预处理后，发现了与血小板中观察到的类似效果，即 (558)Thr 处的 moesin 磷酸化显着增加，并且磷酸化 moesin 与 F-肌动蛋白一起重新分布到细胞质中的一个或多个环状结构，这可能是由于磷酸化 moesin 与 F-肌动蛋白的结合。 (C) 1996 学术出版社
To determine, whether (558)Thr in the carboxyl-terminal domain of moesin is phosphorylated in cells other than platelets, rabbit phosphorylation state-specific antibodies were made to the chemically phosphorylated synthetic hexapeptide KYKpTLR of the moesin sequence, as well as to the unphosphorylated form. The affinity-purified antibody populations were specific for either the phosphorylated or the unmodified peptide conjugated to BSA. Site-specific phosphorylation of moesin is detected in RAW macrophages by Western blot analysis, and immunofluorescence studies demonstrate that phosphorylated moesin is localized in filopodial protrusions. After pretreatment with the phosphatase inhibitor calyculin A, a similar effect to that seen in platelets is found, namely a substantial increase in moesin phosphorylation at (558)Thr and redistribution of phospho-moesin together with F-actin into one or more ring-like structures in the cytoplasm, presumably due to binding of phosphorylated moesin to F-actin. (C) 1996 Academic Press, Inc.