BioProject

Changes in genome structure and gene expression have been documented in both resynthesized and natural allopolyploids that contain two or more divergent genomes. More...

Changes in genome structure and gene expression have been documented in both resynthesized and natural allopolyploids that contain two or more divergent genomes. The underlying mechanisms for rapid and stochastic changes in gene expression are unknown. Arabidopsis suecica is a natural allotetraploid derived from the extant species A. thaliana and A. arenosa. Here we report that reduced DNA methylation in met1-RNAi A. suecica lines altered the expression of ~200 genes encoding transposons, centromeric and heterochromatic RNAs, and predicted proteins. Reduced DNA methylation occurred frequently in promoter regions of the upregulated genes but not of the repressed genes and led to increased mobility of En/Spm-like transposons in met1-RNAi A. suecica lines. Compared to A. arenosa centromeres, A. thaliana centromeres were hypermethylated, which correlates with higher levels of small RNA accumulation in A. thaliana centromeres than that in A. arenosa centromeres. Derepression of the genes examined was primarily derived from A. thaliana subgenome, and A. arenosa genes are less affected by methylation defects. Moreover, non-CG (CC) methylation in the promoter region of A. thaliana At2g23810 was maintained in resynthesized allotetraploids, and the methylation spread within the promoter region in natural A. suecica, leading to silencing of At2g23810, which was demethylated and reactivated in met1-RNAi A. suecica lines. We suggest that a subset of A. thaliana and A. arenosa genes are differentially methylated in natural allopolyploids, and some A. thaliana genes including centromeres are subjected to transcriptional repression and genome-specific RNA-mediated DNA methylation in Arabidopsis allopolyploids. Keywords: gene expression in reduced DNA methylation lines in Arabidopsis allotetraploids Overall design: Spotted oligo-gene microarrays were used for genome-wide gene expression assays. A total of 8 slides were used for 2 biological and 4 technical replications. Each dye-swap (technical replication) consists of two slides. One slides was hybridized with an equal amount of Cy3-labled A. suecica control cDNA and Cy5-labeled met1-RNAi cDNA as probes, and another with the same two cDNA samples labeled in a reverse Cy3- and Cy5-combination. We used 500 ng of mRNA in each labeling reaction using Cy3- or Cy5-dCTP (Amersham Biosciences). The dye-swap experiment was replicated using an independently isolated RNA sample. Raw data were collected using Genepix Pro4.1 after the slides were scanned using Genepix 4000B (Axon Instruments/Molecular Devices, Sunnyvale, CA). The data were processed using a lowess function to remove non-linear components and analyzed using a linear model. This linear model was employed to partition variation in the observed data relative to technical and biological variation. We selected the genes that were differently expressed at a statistically significant level (FDR, a=0.05) under both common variance and per-gene variance. The microarray data were deposited in GEO (/geo/) and an accession number is being assigned. Functional categories of up- and down-regulated genes were classified using (ftp://ftp.arabidopsis.org/home/tair/Ontologies/Gene_Ontology/OLD/ATH_GO_GOSLIM.20070317.txt) (TAIR release on 17 March 2007) and compared using Venn diagrams. The differentially expressed genes (identities) in met1-RNAi lines compared to A. suecica control were mapped to oligonucleotide sequences using Perl scripts. The oligos were mapped to genomic coordinates using high (red) and low (blue) gradients corresponding to gene densities. Vertical lines above and below the chromosomes showed up- and down-regulation, respectively, and the length was proportional to the logarithm-fold changes in differential gene expression. Less...