Dynamics of DNA Methylation and Its Functions in Plant Growth and Development

Abstract

Epigenetic modifications in DNA bases and histone proteins play important roles in the regulation of gene expression and genome stability. Chemical modification of DNA base (e.g., addition of a methyl group at the fifth carbon of cytosine residue) switches on/off the gene expression during developmental process and environmental stresses. The dynamics of DNA base methylation depends mainly on the activities of the writer/eraser guided by non-coding RNA (ncRNA) and regulated by the developmental/environmental cues. De novo DNA methylation and active demethylation activities control the methylation level and regulate the gene expression. Identification of ncRNA involved in de novo DNA methylation, increased DNA methylation proteins guiding DNA demethylase, and methylation monitoring sequence that helps maintaining a balance between DNA methylation and demethylation is the recent developments that may resolve some of the enigmas. Such discoveries provide a better understanding of the dynamics/functions of DNA base methylation and epigenetic regulation of growth, development, and stress tolerance in crop plants. Identification of epigenetic pathways in animals, their existence/orthologs in plants, and functional validation might improve future strategies for epigenome editing toward climate-resilient, sustainable agriculture in this era of global climate change. The present review discusses the dynamics of DNA methylation (cytosine/adenine) in plants, its functions in regulating gene expression under abiotic/biotic stresses, developmental processes, and genome stability.

Keywords: 5-methylcytosine; DNA methylation; DNA modification; N6-methyladenine; environmental stress; epigenetics; gene regulation; plant growth.

Figure 1

Diagrammatic representation of the RNA-directed DNA methylation (RdDM) pathway. According to the canonical RdDM pathway, noncoding P4 RNAs are produced by RNA polymerase IV (Pol IV). SHH1 binds to dimethylated histone H3K9me2 and helps to recruit Pol IV at RdDM locus. (Path 1): P4 RNAs get converted into double-stranded RNAs (dsRNAs) by RDR2, which get cleaved into 24 nucleotide (nt) siRNAs by DICER-like protein 2 (DCL2),ss DCL3, and DCL4. These siRNAs bound with Argonaute 4 or AGO6 participate in the RdDM. (Path 2): Methylation of RdDM loci in dcl1-dcl2-dcl3-dcl4 mutant suggests the existence of DCL-independent RdDM. (Path 3): POL II produces 24 nt siRNAs with the help of DCL3 and scaffold RNAs at some of the RdDM loci. (Path 4): For some active transposons, mRNAs get converted into dsRNAs and get cleaved into 21 nt siRNAs by DCL2, DCL4 through RDR6–RdDM pathway. Involved in de novo (IDN)–IDN2 Paralog (IDP) complex and RNA-binding proteins RRP6-like 1 (RRP6L1) interact with a chromatin-remodeling complex Switch/Sucrose Nonfermenting (SWI/SNF) to facilitate retention of nascent Pol V-transcribed RNA. m, methylcytosine. (Redrawn from Zhang et al., 2018a).

Figure 2

Dynamics of DNA methylation in plants. De novo DNA methylation occurs in all (CG, CHG, and CHH; where H=A, C, or T) cytosine contexts. After replication of DNA, methylation in the CG context is maintained by methyltransferase 1 (MET1), while methylation in CHG context is maintained by chromomethylase 2 (CMT2) or CMT3, and methylation in CHH context is maintained by CMT2 or by DRM2 via RdDM pathway. Methylated CHG (^mCHG) attracts histone H3 lysine 9 (H3K9)-specific suppressor of variegation 3-9 homolog protein 4 (SUVH4), SUVH5, and SUVH6 and generates dimethylated H3K9 (H3K9me2), which enables CMT2 and CMT3 (A). Methylation of methylation monitoring sequence (MEMS), also known as “methylstat” present in the promoter of the Repressor of silencing 1 (Ros1) is necessary for transcription of the Ros1 gene. Cytosine methylation at MEMS is controlled by MET1/RdDM and Ros1 itself. This helps to sense/monitor the level of methylation and regulate DNA (de)methylation homeostasis (B). CH₃, methyl group, Me/m, methylation (Redrawn from Zhang et al., 2018a).

Figure 3

Dynamics of adenine methylation. Adenine (A) gets methylated by the addition of CH₃ (methyl) group at N⁶ position by DNA adenine methyltransferases 1, the writer, generating N⁶-methyladenine (6-mA). SeqA protein, the reader, specifically binds to hemimethylated 6-mA DNA. The 6-mA gets hydroxylated (–OH) by AlkB oxidase to N⁶-hydroxymethylcytosine (6-hmA). Due to the erasers like DNA 6-mA demethylase (DMAD) or N⁶-methyladenine demethylase-1 (NMAD1), 6-mA gets deaminated back to Adenine. 6-hmA gets demethylated indirectly to Adenine by AlkB oxidase via N⁶-hydroxymethyl adenine (6-hmA) and N⁶-formyladenine (6-fA). Adenine may also get methyl adduct to N¹-methyladenine (1-mA) by environmental/endogenous alkylating agents. Similarly, 1-mA may also get demethylated indirectly by AlkB oxidase to Adenine via N¹-hydroxymethyl adenine (1-hmA). AlkB is one of the prototypic oxidative dealkylation DNA repair enzymes/dioxygenases involved in the removal of alkyl adducts from DNA base by oxidative dealkylation (Redrawn from Kumar et al., 2018).

Figure 4

Role of DNA methylation during reproductive development in plants. During male gametogenesis in pollen, the transposons in the vegetative cell are de-silenced due to DNA demethylation by Demeter (DME) and by downregulated expression of Decreased DNA Methylation 1 (DDM1, a chromatin remodeler). Transposon-generated transcripts are converted into siRNAs, which enter into the sperm cells and cause silencing of transposon through DNA methylation. On pollination, one of the sperm cells fertilizes a female gamete (central cell) and forms the endosperm, wherein reinforced CHH methylation at transposons (in the male genome) and DME-mediated global DNA demethylation (in the female genomes) are observed. Another sperm cell fertilizes the egg cell to form an embryo, where reinforced CHH methylation in the male genome but domain rearranged methylase 2 (DRM2) and RNA polymerase V (Pol V) derived RdDM pathway in the female genome are observed.

Figure 5

Role of DNA methylation in stress tolerance and epigenetic stress memory in plants. Abiotic stresses may alter the expression of the stress-responsive genes and cause DNA base modifications. In stressed plants, epigenetic modifications reprogram the epigenetic landscape and some of the epigenetic changes may be inherited (A). Expression of stress-responsive genes may also cause changes in epigenetic modifications. After the stress, during the recovery period, Morpheus Molecule 1 (MOM1) and DDM1 erase the stress-induced epigenetic marks. Inheritance of stress-induced epigenetic marks/de-silencing of genes can be seen in plants repeatedly exposed to the tress due to the dysfunction of DDM1 and MOM1 (B). H3K9me2, dimethylated histone H3 lysine 9 (Modified from Zhang et al., 2018a).