Müller H P, Varmus H E
Department of Microbiology and Immunology, University of California, San Francisco 94143-0502.
EMBO J. 1994 Oct 3;13(19):4704-14. doi: 10.1002/j.1460-2075.1994.tb06794.x.
The choice of retroviral integration sites is strongly influenced by chromatin: integration in vitro occurs more efficiently into nucleosomal DNA than into naked DNA, and a characteristic pattern of preferred insertion sites with a 10 bp periodicity is observed at the outer face of the nucleosomal DNA. At least three features of nucleosomal DNA could be responsible for the creation of these favored sites: the presence of histones, attachment of the DNA to a protein surface, and DNA bending. To test each of these possibilities, we studied integration in vitro with human immunodeficiency virus and murine leukemia virus integrases into four model targets that mimic features of nucleosomal DNA: (i) catabolite activator protein-DNA complexes; (ii) lac repressor-operator complexes; (iii) lac repressor-induced loops; and (iv) intrinsically bent A-tract DNA. We found that bending of the target DNA can create favored integration sites at the outer face of the helix, irrespective of whether the bent DNA is attached to a protein surface. Our findings offer an explanation for the preferred usage of nucleosomes as integration targets. In addition, they suggest that bending of the target DNA might be an intrinsic feature of the integration reaction.
体外整合在核小体DNA中比在裸露DNA中更有效，并且在核小体DNA的外表面观察到具有10 bp周期性的优选插入位点的特征模式。核小体DNA的至少三个特征可能导致这些有利位点的产生：组蛋白的存在、DNA与蛋白质表面的附着以及DNA弯曲。为了测试每种可能性，我们研究了用人类免疫缺陷病毒和鼠白血病病毒整合酶体外整合到模拟核小体DNA特征的四个模型靶标中：（i）分解代谢物激活蛋白-DNA复合物；（ii）乳糖阻遏物-操纵子复合物；（iii）乳糖阻遏物诱导的环；和（iv）内在弯曲的A-序列DNA。我们发现靶DNA的弯曲可以在螺旋的外表面产生有利的整合位点，而不管弯曲的DNA是否附着在蛋白质表面。我们的发现为核小体作为整合靶标的优选使用提供了解释。此外，它们表明靶DNA的弯曲可能是整合反应的一个内在特征。
