Nuclear Dynamics inArabidopsis thaliana

Abstract

The nucleus is a definitive feature of eukaryotic cells, comprising twin bilamellar membranes, the inner and outer nuclear membranes, which separate the nucleoplasmic and cytoplasmic compartments. Nuclear pores, complex macromolecular assemblies that connect the two membranes, mediate communication between these compartments. To explore the morphology, topology, and dynamics of nuclei within living plant cells, we have developed a novel method of confocal laser scanning fluorescence microscopy under time-lapse conditions. This is used for the examination of the transgenic expression in Arabidopsis thaliana of a chimeric protein, comprising the GFP (Green-Fluorescent Protein of Aequorea victoria) translationally fused to an effective nuclear localization signal (NLS) and to ␤-glucuronidase (GUS) from E. coli. This large protein is targeted to the nucleus and accumulates exclusively within the nucleoplasm.This article provides online access to movies that illustrate the remarkable and unusual properties displayed by the nuclei, including polymorphic shape changes and rapid, longdistance, intracellular movement. Movement is mediated by actin but not by tubulin; it therefore appears distinct from mechanisms of nuclear positioning and migration that have been reported for eukaryotes. The GFP-based assay is simple and of general applicability. It will be interesting to establish whether the novel type of dynamic behavior reported here, for higher plants, is observed in other eukaryotic organisms. INTRODUCTIONA definitive feature of eukaryotic cells is the nucleus, first described in stamen cells of Tradescantia by Robert Brown in 1831, which is found in all cell types at some stage of their development. The nucleus comprises twin bilamellar membranes, the inner and outer nuclear membranes, which serve to separate the nuclear contents, the nucleoplasm, from the cytoplasm. Communication between these compartments is mediated by nuclear pores, complex macromolecular assemblies that connect the two membranes.All of our information concerning nuclear morphology, topology, and dynamics comes from the use of various forms of microscopy. The major components of the nucleoplasm, including proteins, RNA, and DNA, do not absorb in the visible part of the spectrum. Thus, nuclei are nearly translucent and nonfluorescent. This impedes the observation of nuclei within living cells with standard bright-field or fluorescence microscopy. Although other forms of light microscopy can be used to examine nuclei, particularly phase contrast and differential interference contrast microscopy, which rely on optical properties other than absorbance and fluorescence, this is practical only in nonpigmented cells and in tissues having simple three-dimensional organization.We have developed an alternative method for observation of nuclei in living plant cells that uses confocal laser scanning fluorescence microscopy (Chytilova et al., 1999). Based on the transgenic expression of the Green Fluorescent Protein (GFP) of Aequorea victoria, it involves ...

Interphase Chromosome Arrangement in Arabidopsis thaliana Is Similar in Differentiated and Meristematic Tissues and Shows a Transient Mirror Symmetry After Nuclear Division

“…Second, to distinguish between actual mirror-symmetry and possible rotation of nuclei (Chytilova et al 2000), 10 adjacent BACs from the AT3 top arm were used as a reference point in addition to the AT2 arm-specific FISH probes. A mirror-symmetric positional arrangement of the homologous AT2 arm territories and the 10 adjacent BACs from AT3 was observed in all six investigated pairs of sister cells derived immediately from initial cells (two pairs shown in Figure 4, A and B, sister cells s1, s2, s3, and s4).…”

How plants LINC the SUN to KASH

“…Interrupting this nuclear positioning by depolymerizing F-actin or trapping the nucleus with a laser beam prohibits root hair growth. 42,43 Similarly, in Arabidopsis trichomes-branched, single-cell leaf hairs-the nucleus migrates to a position close to the first branch point. 44 Asymmetric cell divisions play an important role in plant development.…”

‘Poking’ microtubules bring about nuclear wriggling to position nuclei

“…The actin network has been known to function actively in repositioning the nuclei in the root hairs of Arabidopsis. Drugs that depolymerize the actin cytoskeleton abolish the process, whereas drugs that disrupt the MTs possess no effect (Chytilova et al, 2000). In the budding yeast both the actin filaments and the MTs participate in proper localization of the nuclei and the spindle apparatus to ensure normal cell division (Palmer et al, 1992;Bloom, 2001;).…”