Targeting of Ikaros to pericentromeric heterochromatin by direct DNA binding
Targeting of Ikaros to pericentromeric heterochromatin by direct DNA binding
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DOI：
10.1101/gad.816400
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发表时间：
2000-09-01
影响因子：
10.5
通讯作者：
Smale, ST
中科院分区：
文献类型：
--
作者：
Cobb, BS;Morales-Alcelay, S;Smale, ST
Ikaros是一种序列特异性DNA结合蛋白，对淋巴细胞发育至关重要。关于Ikaros的分子功能知之甚少，尽管最近的研究结果导致了这样的假设，即它招募注定用于遗传失活的基因到含有近着丝粒异染色质的病灶。为了进一步了解Ikaros的功能，我们研究了它靶向着丝粒病灶的机制。在3 T3成纤维细胞中异位表达后观察到Ikaros的有效靶向，表明不需要淋巴细胞特异性蛋白和淋巴核结构。着丝粒周围靶向不是由与Mi-2重塑因子的相互作用引起的，因为在3 T3细胞中只有小百分比的Mi-2定位于着丝粒病灶。相反，靶向依赖于Ikaros的氨基末端DNA结合锌指结构域和羧基末端二聚化结构域。羧基末端结构域仅为同源二聚化所需，因为当该结构域被亮氨酸拉链取代时，靶向恢复。令人惊讶的是，一个详细的氨基末端结构域的取代突变体分析揭示了DNA结合和近着丝粒靶向之间的密切相关性。这些结果表明，DNA结合是必不可少的pericentromeric本地化的Ikaros，可能与存在的Ikaros结合位点内着丝粒DNA重复。与此目标机制一致的Ikaros的功能模型进行了讨论。
Ikaros is a sequence-specific DNA-binding protein that is essential for lymphocyte development. Little is known about the molecular function of Ikaros, although recent results have led to the hypothesis that it recruits genes destined for heritable inactivation to foci containing pericentromeric heterochromatin. To gain further insight into the functions of Ikaros, we have examined the mechanism by which it is targeted to centromeric foci. Efficient targeting of Ikaros was observed upon ectopic expression in 3T3 fibroblasts, demonstrating that lymphocyte-specific proteins and a lymphoid nuclear architecture are not required. Pericentromeric targeting did not result from an interaction with the Mi-2 remodeling factor, as only a small percentage of Mi-2 localized to centromeric foci in 3T3 cells. Rather, targeting was dependent on the amino-terminal DNA-binding zinc finger domain and carboxy-terminal dimerization domain of Ikaros. The carboxy-terminal domain was required only for homodimerization, as targeting was restored when this domain was replaced with a leucine zipper. Surprisingly, a detailed substitution mutant analysis of the amino-terminal domain revealed a close correlation between DNA-binding and pericentromeric targeting. These results show that DNA binding is essential for the pericentromeric localization of Ikaros, perhaps consistent with the presence of Ikaros binding sites within centromeric DNA repeats. Models for the function of Ikaros that are consistent with this targeting mechanism are discussed.