PERTURBATION OF CHROMATIN STRUCTURE IN THE REGION OF THE ADULT BETA-GLOBIN GENE IN CHICKEN ERYTHROCYTE CHROMATIN
PERTURBATION OF CHROMATIN STRUCTURE IN THE REGION OF THE ADULT BETA-GLOBIN GENE IN CHICKEN ERYTHROCYTE CHROMATIN
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DOI：
10.1016/0022-2836(87)90626-7
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发表时间：
1987-01-05
影响因子：
5.6
通讯作者：
ALLAN, J
中科院分区：
文献类型：
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作者：
CAPLAN, A;KIMURA, T;ALLAN, J
从鸡红细胞核中分离出的含有成人 β-珠蛋白基因和侧翼序列的 EcoRI 染色质片段，相对于相同大小的大量染色质片段，其沉积速率较低。我们表明，通过向天然染色质添加额外的连接组蛋白不能逆转特定的延迟。当通过去除接头组蛋白或降低离子强度来展开染色质片段时，珠蛋白和大块染色质片段之间的差异不再可见。然而，通过将接头组蛋白恢复到耗尽的染色质而获得的重折叠染色质表现出原始的沉降差异。因此，这种差异是由于活性基因上组蛋白八聚体的特殊性质决定了其折叠成高阶结构的程度。使用 CsCl 密度梯度测量蛋白质与 DNA 的比率表明，这并不是由于体外接头组蛋白的差异结合所致。在选择性去除接头组蛋白之前和之后，珠蛋白基因片段和大量染色质片段仅表现出浮力密度的微小差异。此外，我们还表明，通过在球蛋白基因侧翼的 5'' 和 3'' 核酸酶过敏位点消化来切割 EcoRI 片段，会从这些位点之间释放出正常沉积的片段。我们得出的结论是，过敏位点本身是沉降速率降低的原因。非核小体 DNA 片段似乎太长，无法整合到染色质螺线管中，因此在染色质中的单独螺线管元件之间形成间隔物，这可以解释其流体动力学行为。
An EcoRI chromatin fragment containing the adult beta-globin gene and flanking sequences, isolated from chicken erythrocyte nuclei, sediments at a reduced rate relative to bulk chromatin fragments of the same size. We show that the specific retardation cannot be reversed by adding extra linker histones to native chromatin. When the chromatin fragments are unfolded either by removing linker histones or lowering the ionic strength, the difference between globin and bulk chromatin fragments is no longer seen. The refolded chromatin obtained by restoring the linker histones to the depleted chromatin, however, exhibits the original sedimentation difference. This difference is therefore due to a special property of the histone octamers on the active gene that determines the extent of its folding into higher-order structure. That it is not due to the differential binding of linker histones in vitro is shown by measurements of the protein to DNA ratios using CsCl density-gradients. Both before and after selective removal of the linker histones, the globin gene fragment and bulk chromatin fragments exhibit only a marginal difference in buoyant density. In addition, we show that cleavage of the EcoRI fragment by digestion at the 5'' and 3'' nuclease hypersensitive sites flanking the globin gene liberates a fragment from between these sites that sediments normally. We conclude that the hypersensitive sites per se are responsible for the reduction in sedimentation rate. The non-nucleosomal DNA segments appear to be too long to be incorporated into the chromatin solenoid and thus create spacers between separate solenoidal elements in the chromatin, which can account for its hydrodynamic behaviour.