Repair of UV lesions in silenced chromatin provides in vivo evidence for a compact chromatin structure
Repair of UV lesions in silenced chromatin provides in vivo evidence for a compact chromatin structure
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DOI：
10.1074/jbc.m306335200
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发表时间：
2003-09-26
影响因子：
4.8
通讯作者：
Thoma, F
中科院分区：
文献类型：
--
作者：
Livingstone-Zatchej, M;Marcionelli, R;Thoma, F
酵母中靠近端粒的基因被含有SIR蛋白的异染色质样结构沉默。为了研究沉默是否也影响DNA修复，我们研究了通过光解酶和核苷酸切除修复(NER)去除紫外线损伤，这些菌株含有从端粒插入2千碱基的URA3基因。在sir3Delta突变体中，URA3在转录上是活跃的，在SIR3细胞中部分沉默，或因SIR3的过度表达或RPD3的缺失而完全沉默。激活的URA3在两条通路上都显示出有效的修复作用。光解酶对启动子和3‘端的快速修复反映了一种非核小体结构。部分沉默对光解酶没有显著影响，但减少了NER的修复，表明两种修复反应的可及性不同。完全沉默抑制了编码区以及启动子和3‘端的NER和光解酶。传统的核酸酶足迹分析显示，在部分沉默的条件下，启动子近端的核小体发生了细微的变化，但在沉默的染色质中，染色质发生了显着的重组，延伸到整个基因。因此，这两个修复系统都对与沉默相关的染色质变化敏感，并为异染色质的紧凑结构提供了直接证据。
Genes positioned close to telomeres in yeast are silenced by a heterochromatin-like structure containing Sir proteins. To investigate whether silencing also affects DNA repair, we studied removal of UV lesions by photolyase and nucleotide excision repair (NER) in strains containing the URA3 gene inserted 2 kilobases from a telomere. URA3 was transcriptionally active in sir3Delta mutants, partially silenced in SIR3 cells, or completely silenced by overexpression of SIR3 or deletion of RPD3. The active URA3 showed efficient repair by both pathways. Fast repair of the promoter and 3' end by photolyase reflected a non-nucleosomal structure. Partial silencing had no remarkable effect on photolyase but reduced repair by NER, indicating differential accessibility for the two repair reactions. Complete silencing inhibits NER and photolyase in the coding region as well as in the promoter and the 3'-end. Conventional nuclease footprinting analyses revealed subtle changes in the promoter proximal nucleosome under partially silenced conditions but a pronounced reorganization of chromatin extending over the whole gene in silenced chromatin. Thus, both repair systems are sensitive to chromatin changes associated with silencing and provide direct evidence for a compact structure of heterochromatin.