Bridger J M, Herrmann H, Münkel C, Lichter P
Organization of Complex Genomes, German Cancer Research Center, Heidelberg, Germany.
J Cell Sci. 1998 May;111 ( Pt 9):1241-53. doi: 10.1242/jcs.111.9.1241.
A number of structural and functional subnuclear compartments have been described, including regions exclusive of chromosomes previously hypothesized to form a reactive nuclear space. We have now explored this accessible nuclear space and interchromosomal nucleoplasmic domains experimentally using Xenopus vimentin engineered to contain a nuclear localization signal (NLS-vimentin). In stably transfected human cells incubated at 37 degrees C, the NLS-vimentin formed a restricted number of intranuclear speckles. At 28 degrees C, the optimal temperature for assembly of the amphibian protein, NLS-vimentin progressively extended with time out from the speckles into strictly orientated intranuclear filamentous arrays. This enabled us to observe the development of a system of interconnecting channel-like areas. Quantitative analysis based on 3-D imaging microscopy revealed that these arrays were localized almost exclusively outside of chromosome territories. During mitosis the filaments disassembled and dispersed throughout the cytoplasm, while in anaphase-telophase the vimentin was recruited back into the nucleus and reassembled into filaments at the chromosome surfaces, in distributions virtually identical to those observed in the previous interphase. The filaments also colocalized with specific nuclear RNAs, coiled bodies and PML bodies, all situated outside of chromosome territories, thereby interlinking these structures. This strongly implies that these nuclear entities coexist in the same interconnected nuclear compartment. The assembling NLS-vimentin is restricted to and can be used to delineate, at least in part, the formerly proposed reticular interchromosomal domain compartment (ICD). The properties of NLS-vimentin make it an excellent tool for performing structural and functional studies on this compartment.
人们已经描述了许多结构和功能上的亚核区室，包括先前假设形成反应性核空间的不含染色体的区域。我们现在利用工程改造后含有核定位信号的非洲爪蟾波形蛋白（NLS-波形蛋白），通过实验探索了这个可及的核空间和染色体间核质结构域。在37℃培养的稳定转染的人类细胞中，NLS-波形蛋白形成数量有限的核内斑点。在28℃（两栖类蛋白组装的最佳温度）时，NLS-波形蛋白随着时间的推移逐渐从斑点处向外延伸，形成严格定向的核内丝状阵列。这使我们能够观察到一个相互连接的通道样区域系统的形成。基于三维成像显微镜的定量分析表明，这些阵列几乎完全位于染色体区域之外。在有丝分裂期间，细丝解体并分散到整个细胞质中，而在后期到末期，波形蛋白被重新募集回细胞核，并在染色体表面重新组装成细丝，其分布与之前间期观察到的几乎相同。这些细丝还与特定的核RNA、卷曲小体和早幼粒细胞白血病蛋白（PML）小体共定位，所有这些都位于染色体区域之外，从而将这些结构相互连接起来。这强烈暗示这些核实体共存于同一个相互连接的核区室中。组装中的NLS-波形蛋白至少部分地局限于先前提出的网状染色体间结构域区室（ICD），并可用于描绘该区域。NLS-波形蛋白的特性使其成为对该区域进行结构和功能研究的优秀工具。
