Modifications of human histone H3 variants during mitosis
Modifications of human histone H3 variants during mitosis
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DOI：
10.1021/bi050906n
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发表时间：
2005-10-04
期刊：
影响因子：
2.9
通讯作者：
Hunt, DF
中科院分区：
文献类型：
--
作者：
Garcia, BA;Barber, CM;Hunt, DF
组蛋白H3的磷酸化是有丝分裂中的标志性事件，并且与染色体浓缩相关。在这里，我们使用固定化金属亲和层析和串联质谱的组合来表征与磷酸化相关的翻译后修饰的组蛋白H3变体的N-末端尾部纯化有丝分裂阻滞的HeLa细胞。在体内观察到的与磷酸化的Ser和Thr相邻的赖氨酸残基上的修饰为“甲基/磷酸”、二元开关假说的存在提供了支持[Fischle，W.，王玉，和Allis，C. D.（2003）Nature 425，475-479]。当邻近的Lys残基被修饰时，用对H3在Ser 10、Ser 28和Thr 3处具有选择性的抗体进行的ELISA显示出降低的活性。当质谱和免疫测定方法一起使用时，为阐明组蛋白编码和鉴定有丝分裂和其他特定细胞事件期间发生的组蛋白翻译后修饰提供了强有力的方法。
Phosphorylation of histone H3 is a hallmark event in mitosis and is associated with chromosome condensation. Here, we use a combination of immobilized metal affinity chromatography and tandem mass spectrometry to characterize post-translational modifications associated with phosphorylation on the N-terminal tails of histone H3 variants purified from mitotically arrested HeLa cells. Modifications observed in vivo on lysine residues adjacent to phosphorylated Ser and Thr provide support for the existence of the "methyl/phos", binary-switch hypothesis [Fischle, W., Wang, Y., and Allis, C. D. (2003) Nature 425, 475-479]. ELISA with antibodies selective for H3 at Ser10, Ser28, and Thr3 show reduced activity when adjacent Lys residues are modified. When used together, mass spectrometry and immunoassay methods provide a powerful approach for elucidation of the histone code and identification of histone post-translational modifications that occur during mitosis and other specific cellular events.