Gene Regulation Group, Centre for Biomolecular Sciences, School of Pharmacy, University of Nottingham, University Park, Nottingham NG7 2RD, United Kingdom.
Biochem Biophys Res Commun. 2010 Jan 1;391(1):1136-41. doi: 10.1016/j.bbrc.2009.12.040. Epub 2009 Dec 16.
The lysine acetyltransferase CREB binding protein (CBP) is required for chromatin modification and transcription at many gene promoters. In fixed cells, a large proportion of CBP colocalises to PML or nuclear bodies. Using live cell imaging, we show here that YFP-tagged CBP expressed in HEK293 cells undergoes gradual accumulation in nuclear bodies, some of which are mobile and migrate towards the nuclear envelope. Deletion of a short lysine-rich domain that contains the major SUMO acceptor sites of CBP abrogated its ability to be SUMO modified, and prevented its association with endogenous SUMO-1/PML speckles in vivo. This SUMO-defective CBP showed enhanced ability to co-activate AML1-mediated transcription. Deletion mapping revealed that the SUMO-modified region was not sufficient for targeting CBP to PML bodies, as C-terminally truncated mutants containing this domain showed a strong reduction in accumulation at PML bodies. Fluorescence recovery after photo-bleaching (FRAP) experiments revealed that YFP-CBPDelta998-1087 had a retarded recovery time in the nucleus, as compared to YFP-CBP. These results indicate that SUMOylation regulates CBP function by influencing its shuttling between nuclear bodies and chromatin microenvironments.
赖氨酸乙酰转移酶 CREB 结合蛋白 (CBP) 是许多基因启动子的染色质修饰和转录所必需的。在固定细胞中，很大一部分 CBP 与 PML 或核体共定位。在这里，我们通过活细胞成像显示，在 HEK293 细胞中表达的 YFP 标记的 CBP 逐渐积累在核体中，其中一些是可移动的，并向核膜迁移。缺失含有 CBP 主要 SUMO 接受位点的短赖氨酸丰富结构域会破坏其 SUMO 修饰的能力，并阻止其与体内的内源性 SUMO-1/PML 斑点结合。这种 SUMO 缺陷型 CBP 显示出增强的 AML1 介导转录的共激活能力。缺失作图显示，SUMO 修饰的区域不足以将 CBP 靶向到 PML 体中，因为含有该结构域的 C 端截断突变体在 PML 体中的积累明显减少。荧光恢复后光漂白 (FRAP) 实验表明，与 YFP-CBP 相比，YFP-CBPDelta998-1087 在核中的恢复时间较慢。这些结果表明，SUMO 化通过影响 CBP 在核体和染色质微环境之间的穿梭来调节其功能。
