Chelsky D, Olson J F, Koshland D E
J Biol Chem. 1987 Mar 25;262(9):4303-9.
Previous work from this laboratory has shown that approximately 24 proteins are reversibly modified by methyl esterification in a mouse lymphoma cell line. Here, we analyze several mouse tissues as well as other mouse, hamster, and human cell lines and find that many protein-methyl esters are ubiquitous while others show apparent tissue specificity. One of the modified proteins is identified by cellular localization and immunological detection as lamin B, a nuclear envelope structural protein which undergoes depolymerization during mitosis. The average stoichiometry of methylation is at least 0.5 methyl groups per lamin B molecule as determined by radioactive incorporation. By immunoblotting, however, demethylation appears to result in a gain of two negative charges suggesting the loss of two neutral methyl esters producing two carboxylic acid groups per molecule. By comparing mitotic and interphase cells, lamin B is found to be demethylated in mitosis while most other methyl esterified proteins show no appreciable cell cycle dependence. In addition to the correlation with cell cycle, it is shown that lamin B does not incorporate radioactive methyl esters in intact mouse brain tissue yet can do so if the cells are lysed. Analysis of lamin B charge by immunoblotting after isoelectric focusing indicates that this protein is fully methylated in brain suggesting that turnover of methyl groups in intact brain tissue is inhibited. We propose that methylation of lamin B may be involved in the control of disassembly and reassembly of the nuclear envelope during mitosis. If this were the case, the apparent lack of methyl group turnover in brain would be consistent with the inability of those cells to divide.
该实验室之前的研究表明，在小鼠淋巴瘤细胞系中约有24种蛋白质可被甲酯化可逆修饰。在此，我们分析了几种小鼠组织以及其他小鼠、仓鼠和人类细胞系，发现许多蛋白质甲酯普遍存在，而其他一些则表现出明显的组织特异性。其中一种被修饰的蛋白质通过细胞定位和免疫检测被鉴定为核纤层蛋白B，它是一种核膜结构蛋白，在有丝分裂期间会发生解聚。通过放射性掺入测定，核纤层蛋白B甲基化的平均化学计量比为每个分子至少0.5个甲基基团。然而，通过免疫印迹法，去甲基化似乎导致增加了两个负电荷，这表明每个分子失去了两个中性甲酯，产生了两个羧酸基团。通过比较有丝分裂期和间期细胞，发现核纤层蛋白B在有丝分裂期去甲基化，而大多数其他甲酯化蛋白质没有明显的细胞周期依赖性。除了与细胞周期的相关性外，研究还表明，完整小鼠脑组织中的核纤层蛋白B不会掺入放射性甲酯，但如果细胞被裂解则可以。等电聚焦后通过免疫印迹法分析核纤层蛋白B的电荷表明，该蛋白在脑中完全甲基化，这表明完整脑组织中甲基基团的周转受到抑制。我们提出，核纤层蛋白B的甲基化可能参与有丝分裂期间核膜的解体和重新组装的控制。如果是这样，脑中明显缺乏甲基基团周转与这些细胞无法分裂是一致的。
