DNA methylation and its effects on gene expression during primary to secondary growth in poplar stems

- DNA methylation and its effects on gene expression during primary to secondary growth in poplar stems.
- Further analysis revealed a perceptible global correlation between 5mC methylation levels of gene bodies and transcript levels but failed to reveal a correlation between 5mC methylation levels of proximal promoter regions and transcript levels.
- Moreover, DNA methylation exhibits tissue specific pat- terns in plants.
- Although DNA methylation is purported to play an important role in wood formation [30, 31], the mechanisms by which.
- DNA methylation alter the expression of xylogenetic genes have not been elucidated.
- The expression levels of genes involved in DNA methylation and demethylation in P.
- trichocarpa stems To determine whether variations in DNA methylation exist among PS, TS, and SS, we first used qRT-PCR to glo- bally scrutinize the expression levels of genes involved in DNA methylation.
- Variations in the expression levels of genes involved in DNA methylation suggest that genomic DNA methylation levels might be different across PS, TS, and SS.
- To investigate the genomics methylation levels of poplar in the stems of different developmen- tal stages, we performed bisulfite sequencing of gen- omic DNA extracted from PS, TS, and SS using the Illumina HiSeq 2500 platform.
- DNA methylation landscapes of P.
- Moreover, the gene-rich regions with few or no TEs exhibited a relatively less methylation levels.
- trichocarpa stems Given the existence of tissue level variation in DNA methylation in the P.
- In PS, TS, and SS, CG and CHG methylation levels were higher than CHH methylation levels in each of the genomic regions mentioned above (Fig.
- In addition, methylation levels in CG, CHG, and CHH contexts were slightly higher in TS and SS than in PS (Fig.
- In contrast, SINEs had the lowest methylation levels in all three contexts and stages of stem development.
- trichocarpa genomes [37], had higher methylation levels than others in the stems of P..
- In addition, the LTR Copia and LTR Gypsy super families had no distinct differences in their methylation levels across PS, SS, and TS, which resembles their relatively invariant methyla- tion levels across seven tissues (vegetative bud, male in- florescence, female catkin, leaf, root, xylem, and phloem) of P.
- Among differ- ent genic regions, the 5’UTR and 3’UTR had much lower methylation levels than other regions.
- promoters and 2 kb downstream regions had higher methylation levels than other regions in all three methylation contexts in PS, TS, and SS (Fig.
- We also found that methylation levels changed during stem development in P.
- In TEs and genic regions, methylation levels of CG, CHG, and CHH contexts were increased in TS and SS compared to PS (Fig.
- 5c and d), and methylation levels in CG and CHG contexts were highest in TS.
- However, the methylation levels in CHH contexts were highest in SS.
- In both CG and CHG contexts, TEs had higher methylation levels than 2 kb upstream and 2 kb downstream regions.
- Additional studies showed that several TE super families, including LINE L1, DNA CMC-EnSpm, DNA hAT-Tag1, and DNA hAT-Tlp100, had higher CG and CHG methylation levels in TEs than their 2 kb upstream and 2 kb down- stream regions (Additional file 8).
- In addition, the LTR Copia and LTR Gypsy super families had no distinct dif- ferences in three CG, CHG, and CHH methylation levels among TEs and their 2 kb upstream and downstream re- gions in all three tissues..
- Moreover, pro- moter regions had higher methylation levels compared with either gene bodies or 2 kb downstream regions in all three stem tissues.
- To compare methylation levels in the three genomic contexts in different genic regions across multiple tissues, multiple comparison testing was conducted.
- Within the promoter regions, there were no significant differences in CG methylation levels among PS, TS, and SS (Additional file 9A).
- There were significant differ- ences in CHG methylation levels between PS and TS and also between PS and SS (Additional file 9B) and significant differences in CHH methylation levels among PS, TS, and SS (Additional file 9C).
- Within the 2 kb down- stream regions, there were significant differences in CG methylation levels between PS and SS, in CHG between.
- 5 DNA methylation patterns in different genic regions.
- a Methylation levels in different types of transposable elements (TEs), including long terminal repeats (LTR), long interspersed nuclear elements (LINE), short interspersed nuclear elements (SINE), and DNA transposons (DNA) in primary stems (PS), transitional stems (TS), and secondary stems (SS).
- b Methylation levels of gene features, including promoter, 2 kb downstream region, and gene body regions with 5 ’ UTR, exon, intron, and 3 ’ UTR regions in stems of poplar.
- c Methylation levels among TE regions and their 2 kb upstream and downstream regions in stems of poplar.
- d Distribution of methylation levels among gene features, including promoter, gene body, and 2 kb downstream regions in stems of poplar.
- The y-axis indicates methylation levels.
- However, despite being statisti- cally significant, the variations observed in methylation levels are limited..
- Relation between DNA methylation and gene expression To investigate the potential influence of gene methyla- tion on gene expression during wood formation, tran- scriptome profiling of PS, TS, and SS was conducted using the same materials used for methylome analysis..
- 6, the four differen- tially expressed groups had different methylation levels in the three methylation contexts among proximal pro- moters, gene bodies, and 2 kb downstream regions.
- As expected, non-expressed genes had the highest CHG and CHH methylation levels in gene body regions and the highest CG, CHG, and CHH methylation levels within 2 kb downstream regions in PS, TS, and SS.
- In contrast, non-expressed genes had the lowest CHH methylation levels and moderate CG and CHG methyla- tion levels in upstream 2 kb promoter regions in PS, TS, and SS.
- Interestingly, regardless of their expression levels, expressed genes had higher CG, but lower CHG and CHH, methylation levels than non-expressed genes in gene bodies in PS, TS, and SS.
- Expressed genes with moderate expression levels had the highest CG methyla- tion levels within gene bodies, and the highest CG and CHG methylation levels in promoter regions in PS, TS,.
- and SS.
- These results suggest that genes with different expression levels correspond to different CG, CHG, and CHH methylation levels in different genic regions..
- genes in the methylated group were further divided into three subgroups: the bottom third were referred to as the low-methylation subgroup (Low), the middle third as the moderate-methylation subgroup (Moderate), and the top third as the high- methylation group (High) based on their methylation levels.
- The pro- portion of genes with the lowest methylation levels within promoters and with the highest methylation levels within 2 kb downstream regions were the lowest among the three methylation levels, no matter which type of methylation contexts they were.
- To further study the relationship between genic methy- lation and gene expression, Spearman correlation analysis was performed between methylation levels in the whole gene frame (bodies ±2 kb flanking regions) and gene expression levels.
- The y-axis indicates methylation levels..
- Methylation levels were partitioned based on gene expression levels.
- Such a rho may indicate that methylation levels in a small fraction of gene bodies have relatively higher correl- ation with their expression levels..
- Enrichment analyses with the DEGs from PS vs TS revealed pathways for phenylpropanoid biosynthesis, phenylalanine metabol- ism, and the biosynthesis of secondary metabolites (Additional file 12B), suggesting that DNA methylation participates in the initial stage of secondary cell wall formation.
- Since transcription factors (TFs) lie at the center of gene regulation, we identified TFs with pronounced alterna- tions in methylation levels.
- 30.39%, and ~ 5.72% methylation levels in CG, CHG, and CHH contexts (Additional file 1), respectively.
- Although methylation levels in poplar stems fell into the ranges described for other angiosperms (CG.
- The relatively low methylation levels in pop- lar stems maybe due, in part, to the fact that the P..
- 37.7%, and ~ 8.5% methylation levels in CG, CHG, and CHH contexts, respectively [47].
- We also found that methylation levels of poplar stems were negatively correlated with gene numbers (Fig.
- Furthermore, CG, CHG, and CHH methylation levels were positively correlated with TE density (Fig.
- For example, methylation levels differ among seven distinct tissue types in P.
- We also found that CG and CHH methylation levels showed significant differences among PS, TS, and SS.
- The methy- lation levels in CG contexts were highest in TS, and the methylation levels in CHH contexts were highest in SS (Fig.
- It has been reported that DNA methylation can re- press gene expression [56].
- However, the relationship between DNA methylation and gene transcription is more nuanced than initially realized.
- In this study, we found that the relationship between DNA methylation and gene expression was complicated by genic regions, methylation contexts, and developmental states.
- However, in our study, we found that the expressed genes in PS, TS, and SS had higher CG but lower CHG and CHH methylation levels in gene bodies than non-expressed genes (Fig.
- In general, we found that, compared to expressed genes, non-expressed genes in PS, TS, and SS had the lowest CHH methyla- tion in promoter regions, the highest CHG and CHH methylation levels in gene bodies, and the highest CG and CHG methylation levels in 2 kb downstream regions (Fig.
- In this study, we found that, regardless of developmental stage (PS, TS, and SS), CHH methylation levels in promoter regions positively corresponded to gene expression levels (Fig.
- However, CHH methylation levels in the promoter regions of active genes were not consistently higher than those of other gene groups, par- ticularly genes with moderate expression levels.
- According to an earlier study, differences in the expression of specific genes with unique methylation pat- terns, rather than relative methylation levels between the two tissue types, plays a critical role in wood biosynthesis [31].
- DNA methylation analysis using qRT-PCR.
- The methylation levels of each of the three methylation types, CG, CHG, and CHH, were calculated in the whole genome, in each chromosome, and in different genomic regions.
- For all genic regions, methylation profiling of gene bodies and 2 kb flanking regions (both sides of the genes) were plot- ted based on the average methylation levels of different sliding windows..
- Spearman correlation analysis was performed to discern statistical relationship between DNA methylation and gene expression within gene bodies and their ±2 kb flanking regions.
- Methylation levels in various transposable elements (TEs) and their 2 kb proximal regions in primary stems (PS), transitional stems (TS), and secondary stems (SS).
- The y-axis represents methylation levels..
- Comparison of methylation levels in different developmental stages: primary stems (PS), transitional stems (TS), and secondary stems (SS).
- The y-axis represents methylation levels.
- The y-axis indicates the methylation levels.
- (A), (B), and (C) represent CG, CHG, and CHH DNA methylation contexts, respectively..
- Genome-wide analysis of Arabidopsis thaliana DNA methylation uncovers an interdependence between methylation and transcription.
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- Gardening the genome: DNA methylation in Arabidopsis thaliana.
- Widespread dynamic DNA methylation in response to biotic stress..
- Control of CpNpG DNA methylation by the KRYPTONITE histone H3 methyltransferase

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