Protein-DNA interactions in soluble telosomes from Saccharomyces cerevisiae.
Protein-DNA interactions in soluble telosomes from Saccharomyces cerevisiae.
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DOI：
10.1093/nar/23.9.1454
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发表时间：
1995-05
影响因子：
14.9
通讯作者：
J. Wright;V. Zakian
中科院分区：
文献类型：
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作者：
J. Wright;V. Zakian
酵母菌的端粒DNA被组织成一种非核染色体染色质结构，称为端粒，可以通过核酸酶消化以可溶性形式从染色体末端释放出来（Wright， J. H., Gottschling， D. E. and Zakian， V. a . (1992) Genes Dev. 6, 197-210）。通过监测分离的端粒DNA片段，利用凝胶迁移位移和硝化纤维素过滤结合试验来研究可溶性端粒的蛋白质-DNA相互作用。在溴化乙啶解离核小体的浓度下，端粒蛋白仍然与端粒DNA相关。与已知的破坏纤毛虫末端结合蛋白与端粒DNA的相互作用或核小体中组蛋白与DNA的相互作用的盐浓度相比，酵母端粒中的蛋白质-DNA相互作用也被低得多的盐浓度破坏。综上所述，这些数据证实了先前发表的核酸酶作图数据，表明端粒在结构上与传统核小体不同。这些数据还表明，酵母不具有类似于在高盐条件下仍与端粒DNA紧密结合的纤毛虫中检测到的端粒结合蛋白。此外，可溶性端粒的凝胶迁移特性可以通过酵母端粒DNA和重组Rap1p在体外形成的复合物来模拟（Wright， J. H., Gottschling， D. E. and Zakian， V. a .(1992)，可溶性酵母端粒的已知成分Rap1p）。Conrad， M. N., Wright， J. H., Wolf， A. J.和Zakian， V. A. (1990) Cell 63, 739-750)，可能是直接与酵母端粒DNA结合的主要结构蛋白。
Telomeric DNA in Saccharomyces is organized into a non-nucleosomal chromatin structure called the telosome that can be released from chromosome ends in soluble form by nuclease digestion (Wright, J. H., Gottschling, D. E. and Zakian, V. A. (1992) Genes Dev. 6, 197-210). The protein-DNA interactions of soluble telosomes were investigated by monitoring isolated telomeric DNA fragments for the retention of bound protein using both gel mobility shift and nitrocellulose filter-binding assays. Telosomal proteins remained associated with telomeric DNA at concentrations of ethidium bromide that dissociated nucleosomes. The protein-DNA interactions in the yeast telosome were also disrupted by much lower salt concentrations than those known to disrupt either the interactions of ciliate terminus-binding proteins with telomeric DNA or the interactions of histones with DNA in nucleosomes. Taken together, these data corroborate previously published nuclease mapping data indicating that telosomes are distinct in structure from conventional nucleosomes. These data also indicate that yeast do not possess telomere binding proteins similar to those detected in ciliates that remain tightly bound to telomeric DNA even in high salt. In addition, the characteristic gel mobility shift of soluble telosomes could be mimicked by complexes formed in vitro with yeast telomeric DNA and recombinant Rap1p suggesting that Rap1p, a known component of soluble yeast telosomes (Wright, J. H., Gottschling, D. E. and Zakian, V. A. (1992) Genes Dev. 6, 197-210; Conrad, M. N., Wright, J. H., Wolf, A. J. and Zakian, V. A. (1990) Cell 63, 739-750), is likely to be the major structural protein bound directly to yeast telomeric DNA.