Characterization of a nuclear compartment shared by nuclear bodies applying ectopic protein expression and correlative light and electron microscopy

To investigate the accessibility of interphase nuclei for nuclear body-sized particles, we analyzed in cultured cells from human origin by correlative fluorescence and electron microscopy (EM) the bundle-formation of Xenopus-vimentin targeted to the nucleus via a nuclear localization signal (NLS). Moreover, we investigated the spatial relationship of speckles, Cajal bodies, and crystalline particles formed by Mx1 fused to yellow fluorescent protein (YFP), with respect to these bundle arrays. At 37 degrees C, the nucleus-targeted, temperature-sensitive Xenopus vimentin was deposited in focal accumulations. Upon shift to 28 degrees C, polymerization was induced and filament arrays became visible. Within 2 h after temperature shift, arrays were found to be composed of filaments loosely embedded in the nucleoplasm. The filaments were restricted to limited areas of the nucleus between focal accumulations. Upon incubation at 28 degrees C for several hours, NLS vimentin filaments formed bundles looping throughout the nuclei. Speckles and Cajal bodies frequently localized in direct neighborhood to vimentin bundles. Similarly, small crystalline particles formed by YFP-tagged Mx1 also located next to vimentin bundles. Taking into account that nuclear targeted vimentin locates in the interchromosomal domain (ICD), we conclude that nuclear body-sized particles share a common nuclear space which is controlled by higher order chromatin organization.