Bass H W, Nagar S, Hanley-Bowdoin L, Robertson D
Department of Biological Science, Florida State University, Tallahassee, FL 32306-4370, USA.
J Cell Sci. 2000 Apr;113 ( Pt 7):1149-60. doi: 10.1242/jcs.113.7.1149.
Tomato golden mosaic virus (TGMV) is a geminivirus that replicates its single-stranded DNA genome through double-stranded DNA intermediates in nuclei of differentiated plant cells using host replication machinery. We analyzed the distribution of viral and plant DNA in nuclei of infected leaves using fluorescence in situ hybridization (FISH). TGMV-infected nuclei showed up to a sixfold increase in total volume and displayed a variety of viral DNA accumulation patterns. The most striking viral DNA patterns were bright, discrete intranuclear compartments, but diffuse nuclear localization was also observed. Quantitative and spatial measurements of high resolution 3-dimensional image data revealed that these compartments accounted for 1-18% of the total nuclear volume or 2-45% of the total nuclear FISH signals. In contrast, plant DNA was concentrated around the nuclear periphery. In a significant number of nuclei, the peripheral chromatin was organized as condensed prophase-like fibers. A combination of FISH analysis and indirect immunofluorescence with viral coat protein antibodies revealed that TGMV virions are associated with the viral DNA compartments. However, the coat protein antibodies failed to cross react with some large viral DNA inclusions, suggesting that encapsidation may occur after significant viral DNA accumulation. Infection by a TGMV mutant with a defective coat protein open reading frame resulted in fewer and smaller viral DNA-containing compartments. Nevertheless, nuclei infected with the mutant virus increased in size and in some cases showed chromosome condensation. Together, these results established that geminivirus infection alters nuclear architecture and can induce plant chromatin condensation characteristic of cells arrested in early mitosis.
番茄金色花叶病毒（TGMV）是一种双生病毒，它利用宿主的复制机制，通过双链DNA中间体在分化的植物细胞核中复制其单链DNA基因组。我们使用荧光原位杂交（FISH）分析了感染叶片细胞核中病毒DNA和植物DNA的分布。受TGMV感染的细胞核总体积增加了多达六倍，并呈现出多种病毒DNA积累模式。最显著的病毒DNA模式是明亮、离散的核内区室，但也观察到了弥散的核定位。对高分辨率三维图像数据的定量和空间测量表明，这些区室占细胞核总体积的1 - 18%或占细胞核FISH信号总量的2 - 45%。相比之下，植物DNA集中在核周边。在大量细胞核中，周边染色质被组织成类似前期凝聚的纤维。FISH分析与病毒外壳蛋白抗体间接免疫荧光相结合的结果表明，TGMV病毒粒子与病毒DNA区室相关。然而，外壳蛋白抗体未能与一些大的病毒DNA包涵体发生交叉反应，这表明衣壳化可能在病毒DNA大量积累之后发生。用外壳蛋白开放阅读框有缺陷的TGMV突变体感染导致含病毒DNA的区室数量减少且体积变小。尽管如此，感染突变病毒的细胞核大小增加，在某些情况下还出现了染色体凝聚。总之，这些结果表明双生病毒感染会改变核结构，并能诱导植物染色质凝聚，这是早期有丝分裂停滞细胞的特征。
