Limits...
Small RNA-directed epigenetic natural variation in Arabidopsis thaliana.

Zhai J, Liu J, Liu B, Li P, Meyers BC, Chen X, Cao X - PLoS Genet. (2008)

Bottom Line: Here, we report that, in the model plant Arabidopsis thaliana, a cluster of approximately 24 nt siRNAs found at high levels in the ecotype Landsberg erecta (Ler) could direct DNA methylation and heterochromatinization at a hAT element adjacent to the promoter of FLOWERING LOCUS C (FLC), a major repressor of flowering, whereas the same hAT element in ecotype Columbia (Col) with almost identical DNA sequence, generates a set of low abundance siRNAs that do not direct these activities.We have called this hAT element MPF for Methylated region near Promoter of FLC, although de novo methylation triggered by an inverted repeat transgene at this region in Col does not alter its FLC expression.A genome-wide comparison of Ler and Col small RNAs identified at least 68 loci matched by a significant level of approximately 24 nt siRNAs present specifically in Ler but not Col, where nearly half of the loci are related to repeat or TE sequences.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Plant Genomics, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China.

ABSTRACT
Progress in epigenetics has revealed mechanisms that can heritably regulate gene function independent of genetic alterations. Nevertheless, little is known about the role of epigenetics in evolution. This is due in part to scant data on epigenetic variation among natural populations. In plants, small interfering RNA (siRNA) is involved in both the initiation and maintenance of gene silencing by directing DNA methylation and/or histone methylation. Here, we report that, in the model plant Arabidopsis thaliana, a cluster of approximately 24 nt siRNAs found at high levels in the ecotype Landsberg erecta (Ler) could direct DNA methylation and heterochromatinization at a hAT element adjacent to the promoter of FLOWERING LOCUS C (FLC), a major repressor of flowering, whereas the same hAT element in ecotype Columbia (Col) with almost identical DNA sequence, generates a set of low abundance siRNAs that do not direct these activities. We have called this hAT element MPF for Methylated region near Promoter of FLC, although de novo methylation triggered by an inverted repeat transgene at this region in Col does not alter its FLC expression. DNA methylation of the Ler allele MPF is dependent on genes in known silencing pathways, and such methylation is transmissible to Col by genetic crosses, although with varying degrees of penetrance. A genome-wide comparison of Ler and Col small RNAs identified at least 68 loci matched by a significant level of approximately 24 nt siRNAs present specifically in Ler but not Col, where nearly half of the loci are related to repeat or TE sequences. Methylation analysis revealed that 88% of the examined loci (37 out of 42) were specifically methylated in Ler but not Col, suggesting that small RNA can direct epigenetic differences between two closely related Arabidopsis ecotypes.

Show MeSH
DNA Methylation Analysis in the F1 Heterozygous Plants from the Reciprocal Crosses between Col and Ler.(A) Bisulfite sequencing analysis at MPF (B1 region, see Figure 2) of four heterozygous lines from both the crosses of Col♀×Ler♂ and Ler♀×Col♂. SNPs at MPF between Col and Ler (see Figure S1) were used to distinguish the Col- and Ler-derived sequences from the heterozygous plants. “n” indicates the number of sequenced clones. The DNA methylation status was further confirmed by real-time McrBC-PCR using the McrBC non-digested (white bar) and digested (black bar) DNA from the heterozygotes (same to the DNA samples used in bisulfite sequencing). (B) Real-time McrBC-PCR analysis in 24 more lines from each direction of the crosses to test their methylation status at MPF.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2289841&req=5

pgen-1000056-g005: DNA Methylation Analysis in the F1 Heterozygous Plants from the Reciprocal Crosses between Col and Ler.(A) Bisulfite sequencing analysis at MPF (B1 region, see Figure 2) of four heterozygous lines from both the crosses of Col♀×Ler♂ and Ler♀×Col♂. SNPs at MPF between Col and Ler (see Figure S1) were used to distinguish the Col- and Ler-derived sequences from the heterozygous plants. “n” indicates the number of sequenced clones. The DNA methylation status was further confirmed by real-time McrBC-PCR using the McrBC non-digested (white bar) and digested (black bar) DNA from the heterozygotes (same to the DNA samples used in bisulfite sequencing). (B) Real-time McrBC-PCR analysis in 24 more lines from each direction of the crosses to test their methylation status at MPF.

Mentions: In paramutation, the silenced paramutagenic lines are able to confer the active state of the paramutable lines, and make them become paramutagenic [36]. To test whether the methylated state at MPF in Ler is transmissible, we performed bisulfite sequencing to investigate the DNA methylation status in four F1 lines from the crosses of both Col ♀×Ler ♂ and Ler ♀×Col ♂, with the single nucleotide polymorphisms (SNPs) at MPF (Figure S1) used to distinguish the Col and Ler derived sequencing results (Figure 5A). In addition, twenty-four more lines from reciprocal crosses were tested for their MPF methylation by real-time McrBC-PCR (Figure 5B). These experiments revealed extensive diversity in the methylation status of MPF in each individual line in the F1 generation. This diversity could be summarized in the following way: 1) in some lines, the MPF-siRNAs from Ler are able to trigger the de novo methylation at Col-derived MPF; 2) in some other lines, not only the Col-derived MPF remains unmethylated, the Ler-derived MPF could even lose its methylation; 3) there are also cases in which the Ler-derived MPF remains methylated and Col-derived MPF remains unmethylated, just like their ancestors; therefore the MPF is semi-methylated in the whole plant.


Small RNA-directed epigenetic natural variation in Arabidopsis thaliana.

Zhai J, Liu J, Liu B, Li P, Meyers BC, Chen X, Cao X - PLoS Genet. (2008)

DNA Methylation Analysis in the F1 Heterozygous Plants from the Reciprocal Crosses between Col and Ler.(A) Bisulfite sequencing analysis at MPF (B1 region, see Figure 2) of four heterozygous lines from both the crosses of Col♀×Ler♂ and Ler♀×Col♂. SNPs at MPF between Col and Ler (see Figure S1) were used to distinguish the Col- and Ler-derived sequences from the heterozygous plants. “n” indicates the number of sequenced clones. The DNA methylation status was further confirmed by real-time McrBC-PCR using the McrBC non-digested (white bar) and digested (black bar) DNA from the heterozygotes (same to the DNA samples used in bisulfite sequencing). (B) Real-time McrBC-PCR analysis in 24 more lines from each direction of the crosses to test their methylation status at MPF.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2289841&req=5

pgen-1000056-g005: DNA Methylation Analysis in the F1 Heterozygous Plants from the Reciprocal Crosses between Col and Ler.(A) Bisulfite sequencing analysis at MPF (B1 region, see Figure 2) of four heterozygous lines from both the crosses of Col♀×Ler♂ and Ler♀×Col♂. SNPs at MPF between Col and Ler (see Figure S1) were used to distinguish the Col- and Ler-derived sequences from the heterozygous plants. “n” indicates the number of sequenced clones. The DNA methylation status was further confirmed by real-time McrBC-PCR using the McrBC non-digested (white bar) and digested (black bar) DNA from the heterozygotes (same to the DNA samples used in bisulfite sequencing). (B) Real-time McrBC-PCR analysis in 24 more lines from each direction of the crosses to test their methylation status at MPF.
Mentions: In paramutation, the silenced paramutagenic lines are able to confer the active state of the paramutable lines, and make them become paramutagenic [36]. To test whether the methylated state at MPF in Ler is transmissible, we performed bisulfite sequencing to investigate the DNA methylation status in four F1 lines from the crosses of both Col ♀×Ler ♂ and Ler ♀×Col ♂, with the single nucleotide polymorphisms (SNPs) at MPF (Figure S1) used to distinguish the Col and Ler derived sequencing results (Figure 5A). In addition, twenty-four more lines from reciprocal crosses were tested for their MPF methylation by real-time McrBC-PCR (Figure 5B). These experiments revealed extensive diversity in the methylation status of MPF in each individual line in the F1 generation. This diversity could be summarized in the following way: 1) in some lines, the MPF-siRNAs from Ler are able to trigger the de novo methylation at Col-derived MPF; 2) in some other lines, not only the Col-derived MPF remains unmethylated, the Ler-derived MPF could even lose its methylation; 3) there are also cases in which the Ler-derived MPF remains methylated and Col-derived MPF remains unmethylated, just like their ancestors; therefore the MPF is semi-methylated in the whole plant.

Bottom Line: Here, we report that, in the model plant Arabidopsis thaliana, a cluster of approximately 24 nt siRNAs found at high levels in the ecotype Landsberg erecta (Ler) could direct DNA methylation and heterochromatinization at a hAT element adjacent to the promoter of FLOWERING LOCUS C (FLC), a major repressor of flowering, whereas the same hAT element in ecotype Columbia (Col) with almost identical DNA sequence, generates a set of low abundance siRNAs that do not direct these activities.We have called this hAT element MPF for Methylated region near Promoter of FLC, although de novo methylation triggered by an inverted repeat transgene at this region in Col does not alter its FLC expression.A genome-wide comparison of Ler and Col small RNAs identified at least 68 loci matched by a significant level of approximately 24 nt siRNAs present specifically in Ler but not Col, where nearly half of the loci are related to repeat or TE sequences.

View Article: PubMed Central - PubMed

Affiliation: State Key Laboratory of Plant Genomics, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China.

ABSTRACT
Progress in epigenetics has revealed mechanisms that can heritably regulate gene function independent of genetic alterations. Nevertheless, little is known about the role of epigenetics in evolution. This is due in part to scant data on epigenetic variation among natural populations. In plants, small interfering RNA (siRNA) is involved in both the initiation and maintenance of gene silencing by directing DNA methylation and/or histone methylation. Here, we report that, in the model plant Arabidopsis thaliana, a cluster of approximately 24 nt siRNAs found at high levels in the ecotype Landsberg erecta (Ler) could direct DNA methylation and heterochromatinization at a hAT element adjacent to the promoter of FLOWERING LOCUS C (FLC), a major repressor of flowering, whereas the same hAT element in ecotype Columbia (Col) with almost identical DNA sequence, generates a set of low abundance siRNAs that do not direct these activities. We have called this hAT element MPF for Methylated region near Promoter of FLC, although de novo methylation triggered by an inverted repeat transgene at this region in Col does not alter its FLC expression. DNA methylation of the Ler allele MPF is dependent on genes in known silencing pathways, and such methylation is transmissible to Col by genetic crosses, although with varying degrees of penetrance. A genome-wide comparison of Ler and Col small RNAs identified at least 68 loci matched by a significant level of approximately 24 nt siRNAs present specifically in Ler but not Col, where nearly half of the loci are related to repeat or TE sequences. Methylation analysis revealed that 88% of the examined loci (37 out of 42) were specifically methylated in Ler but not Col, suggesting that small RNA can direct epigenetic differences between two closely related Arabidopsis ecotypes.

Show MeSH