Tandem repetitive transgenes and fluorescent chromatin tags alter local interphase chromosome arrangement in Arabidopsis thaliana. J Cell Sci

University of Geneva, Genève, Geneva, Switzerland
Journal of Cell Science (Impact Factor: 5.43). 09/2005; 118(Pt 16):3751-8. DOI: 10.1242/jcs.02498
Source: PubMed


Fluorescent protein chromatin tagging as achieved by the lac operator/lac repressor system is useful to trace distinct chromatin domains in living eukaryotic nuclei. To interpret the data correctly, it is important to recognize influences of the tagging system on nuclear architecture of the host cells. Within an Arabidopsis line that carries lac operator/lac repressor/GFP transgenes, the transgene loci frequently associate with each other and with heterochromatic chromocenters. Accumulation of tagged fusion protein further enhances the association frequency. Independent experiments with a transgenic plant carrying another multi-copy transgene also revealed, independent of its transcriptional state, unusually high frequencies of association with each other and with heterochromatin. From these results we conclude that the lac operator/lac repressor chromatin tagging system may alter the spatial chromatin organization in the host nuclei (in particular when more than one insertion locus is present) and also that loci of homologous transgenic repeats associate more often with each other and with endogenous heterochromatin than normal euchromatic regions.

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    • "In rice, association occurs between homologous telomeres and centromeres in somatic nuclei (Prieto et al. 2004). Preferential homologous and heterologous associations may also occur at transgenic tandem repetitive sequences, at subtelomeres and pericentromeres in differentiated Arabidopsis nuclei of different endopolyploidy level (Jovtchev et al. 2008, 2011; Pecinka et al. 2005; Watanabe et al. 2005; Schubert et al. 2012). Here, we show that A. thaliana centromeres often associate because only in ∼2–8 % of the 2–8C wild-type nuclei were all ten centromeres separated. "
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    • "To confirm that additional immunosignals result from LacO arrays targeted by the fusion proteins, FISH with centromeric 178-bp repeats (Martinez-Zapater et al. 1986) and with LacO probes (Kato and Lam 2001) was carried out on the same interphase nuclei (Fig. 2a). Previous FISH studies revealed not only frequent pairing of the inserted LacO arrays but also often their association with pericentromeric heterochromatin (in >40 % of nuclei) (Pecinka et al. 2005). Therefore, in our experiments, many nuclei did not show small immunosignals in addition to the centromeric ones. "
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    • "Apparently, not all T-DNA integrations resulted in shortening of chromosomes and telomere seeding. As a control, we analysed the ploidy level and the frequency of chromosome truncation events in a set of plants obtained from agro-infected tetraploid immature embryos using a T-DNA construct without telomere repeats because spontaneous chromosome rearrangements linked with T-DNA integration have been reported for A. thaliana (Aufsatz et al. 2002; Clark and Krysan 2010; Kinoshita et al. 2010; Pecinka et al. 2005). Twenty one independent transgenic plants were inspected regarding putative chromosome truncations. "
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