Spruijt Cornelia G, Bartels Stefanie J J, Brinkman Arie B, Tjeertes Jorrit V, Poser Ina, Stunnenberg Hendrik G, Vermeulen Michiel
Department of Physiological Chemistry and Cancer Genomics Centre, University Medical Center Utrecht, Utrecht, The Netherlands.
Mol Biosyst. 2010 Sep;6(9):1700-6. doi: 10.1039/c004108d. Epub 2010 Jun 4.
The Mi-2/NuRD (NUcleosome Remodeling and histone Deacetylase) chromatin remodeling complex is a large heterogeneous multiprotein complex associated with transcriptional repression. Here we apply a SILAC based quantitative proteomics approach to show that all known Mi-2/NuRD complex subunits co-purify with Cyclin Dependent Kinase 2 Associated Protein1 (CDK2AP1), also known as Deleted in Oral Cancer 1 (DOC-1). DOC-1 displays in vitro binding affinity for methylated DNA as part of the meCpG binding MBD2/NuRD complex. In luciferase reporter assays, DOC-1 is a potent repressor of transcription. Finally, immunofluorescence experiments reveal co-localization between MBD2 and DOC-1 in mouse NIH-3T3 nuclei. Collectively, these results indicate that DOC-1 is a bona fide subunit of the Mi-2/NuRD chromatin remodeling complex.
Mi-2/NuRD（核小体重塑和组蛋白去乙酰化酶）染色质重塑复合体是一种与转录抑制相关的大型异质多蛋白复合体。在此，我们应用基于稳定同位素标记氨基酸在细胞培养中（SILAC）的定量蛋白质组学方法，以表明所有已知的Mi-2/NuRD复合亚基都与细胞周期蛋白依赖性激酶2相关蛋白1（CDK2AP1，也称为口腔癌缺失1（DOC-1））共同纯化。作为甲基化CpG结合MBD2/NuRD复合体的一部分，DOC-1在体外对甲基化DNA显示出结合亲和力。在荧光素酶报告基因检测中，DOC-1是一种有效的转录抑制因子。最后，免疫荧光实验揭示了MBD2和DOC-1在小鼠NIH-3T3细胞核中的共定位。这些结果共同表明，DOC-1是Mi-2/NuRD染色质重塑复合体的一个真正亚基。
