Mapping of Abutilon mosaic geminivirus minichromosomes
Mapping of Abutilon mosaic geminivirus minichromosomes
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DOI：
10.1128/jvi.77.20.10808-10818.2003
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发表时间：
2003-10-01
影响因子：
5.4
通讯作者：
Jeske, H
中科院分区：
文献类型：
--
作者：
Pilartz, M;Jeske, H
苘麻嵌合双链病毒体的单链环状DNA在感染细胞后通过宿主蛋白与双链DNA互补。这种双链 DNA 作为复制和转录的模板，组装到宿主核小体中，产生环状病毒微型染色体。通过使用分离的细胞核，结合核酸酶敏感性测定和连接介导的 PCR 来分析它们的染色质结构，通过印迹杂交评估一维和二维凝胶中的核小体梯和拓扑异构体分布。研究发现，病毒微型染色体存在于至少两个被 11 或 12 个核小体覆盖的确定结构中，从而留下可与其他宿主因子相互作用的开放间隙。无核小体间隙与基因组 DNA 的两个组件（DNA A 和 DNA B）中的启动子结构和复制起点共定位。核小体以至少两个不同周期的交替阶段定位在整个病毒 DNA 上。单体病毒环状双链DNA的拓扑异构体的分布证实了可变染色质结构的存在，揭示了受感染植物叶片发育过程中不同阶段具有11、12或13个超螺旋转数（对应于各自核小体数量）的分子的最大频率。讨论了可变染色质结构对双生病毒基因调控的作用。
The single-stranded circular DNA of Abutilon mosaic gemimvirions is complemented to double-stranded DNA by host proteins after infecting cells. This double-stranded DNA serves as a template for replication as well as transcription and is assembled into host nucleosomes, yielding circular viral minichromosomes. Their chromatin structure was analyzed by use of isolated nuclei combining nuclease sensitivity assays with ligation-mediated PCR, evaluating nucleosomal ladders and topoisomer distributions in one- and two-dimensional gels by blot hybridization. Viral minichromosomes were found to exist in at least two defined structures covered with 11 or 12 nucleosomes, leaving open gaps accessible for interactions with other host factors. Nucleosome-free gaps were colocalized with promoter structures and the origin of replication in both components of genomic DNA (DNA A and DNA B). Nucleosomes were positioned over the entire viral DNA in at least two alternative phases with different periodicities. The distribution of topoisomers of monomeric viral circular double-stranded DNA confirmed the presence of variable chromatin structures revealing maximum frequencies of molecules with either 11, 12, or 13 superhelical turns (corresponding to respective numbers of nucleosomes) at maximal frequency at different stages during leaf development of infected plants. The role of variable chromatin structures for gene regulation of geminiviruses is discussed.