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Epigenome confrontation triggers immediate reprogramming of DNA methylation and transposon silencing in Arabidopsis thaliana F1 epihybrids - PubMed

  • ️Fri Jan 01 2016

Epigenome confrontation triggers immediate reprogramming of DNA methylation and transposon silencing in Arabidopsis thaliana F1 epihybrids

Mélanie Rigal et al. Proc Natl Acad Sci U S A. 2016.

Abstract

Genes and transposons can exist in variable DNA methylation states, with potentially differential transcription. How these epialleles emerge is poorly understood. Here, we show that crossing an Arabidopsis thaliana plant with a hypomethylated genome and a normally methylated WT individual results, already in the F1 generation, in widespread changes in DNA methylation and transcription patterns. Novel nonparental and heritable epialleles arise at many genic loci, including a locus that itself controls DNA methylation patterns, but with most of the changes affecting pericentromeric transposons. Although a subset of transposons show immediate resilencing, a large number display decreased DNA methylation, which is associated with de novo or enhanced transcriptional activation and can translate into transposon mobilization in the progeny. Our findings reveal that the combination of distinct epigenomes can be viewed as an epigenomic shock, which is characterized by a round of epigenetic variation creating novel patterns of gene and TE regulation.

Keywords: Arabidopsis; DNA methylation; gene silencing; transcription; transposable elements.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Fig. 1.
Fig. 1.

A nonparental IBM1 epiallele in F1 epihybrids. (A) DNA methylation at the large IBM1 intron, as determined by Sanger sequencing of bisulfite-treated DNA. Filled and empty circles indicate methylated and unmethylated cytosines, respectively; the color denotes sequence context (red, CG; blue, CHG; green, CHH). (B) CHG methylation of the large IBM1 intron, as determined by Sanger sequencing of bisulfite-treated DNA. Parental origin of IBM1 alleles in C×m and m×C F1 epihybrid plants was assigned based on presence/absence of CG methylation. (C) DNA methylation at the large IBM1 intron, as determined by CHG methylation-sensitive AluI digestion followed by PCR. For each genotype, PCR amplification was performed on digested (+) and undigested DNA (−). A region unmethylated in the WT and a region deprived of the AluI site were amplified as controls of digestion efficiency and DNA loading, respectively. (D) Quantitative RT-PCR analysis of the accumulation of IBM1 full-length mRNA (IBM1-L) in the indicated genotypes.

Fig. 2.
Fig. 2.

Global DNA methylation changes in F1 epihybrids. (A) Kernel density plots of DNA methylation differences between C×m or m×C epihybrids and the reconstructed F1 (rF1) at CG, CHG, and CHH contexts. (B) Mean methylation difference between C×m and rF1 in 100-kb tiles along chromosome 1 (Chr1). The “m” prefix denotes methyl.

Fig. 3.
Fig. 3.

Genic CHG methylation in epihybrids. (A) Average CHG methylation over TEs and genes containing at least one met1 CHG hypermethylation DMR. (B) Heat-map of CHG methylation levels within met1 CHG hypermethylation DMRs. Rows represent the DMRs, and the columns genotypes. (C) Venn diagram of the overlap between genes containing at least one CHG hypermethylation DMR in met1 and genes containing at least one CHG hypomethylation DMR in F1 epihybrids. (D) Genome-browser views of CHG methylation at the indicated loci. (E) The DNA methylation level at the BNS locus in indicated genotypes was determined by McrBC digestion followed by qPCR. The individual m×C F2 plants analyzed include the ones analyzed in Fig. 1 C and D.

Fig. 4.
Fig. 4.

CG remethylation in epihybrids. (A) Cumulative number of CG hypermethylation DMRs in F1 epihybrids in 100-kb tiles along chromosome 1. (B) Heat-map of CG methylation levels within CG hypermethylation DMRs in F1 epihybrids. (C) Genome-browser view of CG methylation at a representative genomic region displaying CG remethylation in F1 epihybrids. (D) Box plot of CG, CHG, and CHH methylation levels in F1 CG-remethylated DMRs. Values were extracted from one replicate per genotype. (E) Box plot of CG, CHG, and CHH methylation levels in F1 CG-remethylated DMRs and in a random set of sequences of equivalent type and length.

Fig. 5.
Fig. 5.

Hypomethylation at TEs in epihybrids. (A) Average distribution of CG, CHG, and CHH methylation in indicated genotypes over VANDAL elements containing at least one F1 CHH and CHG hypo-DMR. (B) Distribution of CG hypomethylation DMR density over DNA/En-Spm elements in F1 epihybrids. (C) Genome-browser view of CG and CHG methylation at CAC1. Col and met1 parental methylation patterns are shown together with the rF1 profile subtracted from the C×m or the m×C F1 profiles. The dashed boxes show regions with CG and CHG hypomethylation in epihybrids. (D) PCR assay of CAC1 excision in indicated genotypes. Amplification of the “absence” band reflects mobilization of the TE. (E) CHG methylation levels at CHG hypomethylation DMRs that show transgressive behavior in F1 epihybrids. Values were extracted from one replicate per genotype. (F) CHG hypomethylation DMR density over LTR/Gypsy elements in F1 epihybrids.

Fig. 6.
Fig. 6.

RNA expression in F1 epihybrids. (A) RNA expression at loci overlapping with different DMR types in F1 epihybrids relative to MPV. (B) Misregulated genes and TEs in met1 grouped according to their transcription level in epihybrids. Inter., intermediate. Only genes with similar expression in both reciprocal F1 epihybrids are shown. (C) Average distribution of CHG and CHH methylation over TEs activated in met1 and transcribed at a met1-like or higher level in F1 epihybrids. (D) Average number of different DMRs overlapping with TEs grouped according to their transcript level in epihybrids. Inter., intermediate. (E) Genome browser view of mRNA accumulation at ATCOPIA48 (AT1TE66380) in indicated genotypes. RNA levels in cmt3 and suvh4/5/6 are from ref. . (F) DNA methylation levels at the 5′ LTR of ATCOPIA48. (G) Genome-browser view of mRNA accumulation at EVD (AT5TE20395) and ATCOPIA21 (AT5TE65370) elements in indicated genotypes. (H) DNA methylation levels at the 5′ LTRs of EVD and ATCOPIA21 elements.

Fig. 7.
Fig. 7.

The 21-nt sRNA accumulation, mRNA, and DNA methylation levels at ATHILA6A and ATENSPM5 elements. (A) Accumulation of 21-nt sRNAs along ATHILA6A and ATENSPM5 elements in parent (Top) and F1 epihybrids (Bottom). The two ORFs in both TEs are depicted as yellow arrows. (B) Box plots of mRNA accumulation at ATHILA6A (Left) and the two ATENSPM5 ORFs (Right) in indicated genotypes. (C) Average distribution of CG, CHG, and CHH methylation in indicated genotypes over ATHILA6A and ATENSPM5 elements producing 21-nt sRNAs in met1.

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