Arabidopsis Zinc-Finger-Like Protein ASYMMETRIC LEAVES2 (AS2) and Two Nucleolar Proteins Maintain Gene Body DNA Methylation in the Leaf Polarity Gene ETTIN (ARF3)

Plant Cell Physiol. 2018 Jul 1;59(7):1385-1397. doi: 10.1093/pcp/pcy031.

Abstract

Arabidopsis ASYMMETRIC LEAVES2 (AS2) plays a critical role in leaf adaxial-abaxial partitioning by repressing expression of the abaxial-determining gene ETTIN/AUXIN RESPONSE FACTOR3 (ETT/ARF3). We previously reported that six CpG dinucleotides in its exon 6 are thoroughly methylated by METHYLTRASFERASE1, that CpG methylation levels are inversely correlated with ETT/ARF3 transcript levels and that methylation levels at three out of the six CpG dinucleotides are decreased in as2-1. All these imply that AS2 is involved in epigenetic repression of ETT/ARF3 by gene body DNA methylation. The mechanism of the epigenetic repression by AS2, however, is unknown. Here, we tested mutations of NUCLEOLIN1 (NUC1) and RNA HELICASE10 (RH10) encoding nucleolus-localized proteins for the methylation in exon 6 as these mutations enhance the level of ETT/ARF3 transcripts in as2-1. Methylation levels at three specific CpGs were decreased in rh10-1, and two of those three overlapped with those in as2-1. Methylation levels at two specific CpGs were decreased in nuc1-1, and one of those three overlapped with that in as2-1. No site was affected by both rh10-1 and nuc1-1. One specific CpG was unaffected by these mutations. These results imply that the way in which RH10, NUC1 and AS2 are involved in maintaining methylation at five CpGs in exon 6 might be through at least several independent pathways, which might interact with each other. Furthermore, we found that AS2 binds specifically the sequence containing CpGs in exon 1 of ETT/ARF3, and that the binding requires the zinc-finger-like motif in AS2 that is structurally similar to the zinc finger-CxxC domain in vertebrate DNA methyltransferase1.

MeSH terms

  • Arabidopsis / genetics*
  • Arabidopsis / metabolism
  • Arabidopsis Proteins / genetics
  • Arabidopsis Proteins / metabolism*
  • CpG Islands
  • Cytosine / metabolism
  • DEAD-box RNA Helicases / genetics
  • DEAD-box RNA Helicases / metabolism*
  • DNA Methylation*
  • DNA-Binding Proteins / metabolism
  • Exons
  • Gene Expression Regulation, Plant
  • Nuclear Proteins / metabolism
  • Plant Leaves / genetics
  • Plant Leaves / physiology*
  • Protein Domains
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism
  • Transcription Factors / genetics
  • Transcription Factors / metabolism*

Substances

  • AS2 protein, Arabidopsis
  • Arabidopsis Proteins
  • DNA-Binding Proteins
  • ETT protein, Arabidopsis
  • NUC-L1 protein, Arabidopsis
  • Nuclear Proteins
  • RNA-Binding Proteins
  • Transcription Factors
  • Cytosine
  • DEAD-box RNA Helicases
  • RH10 protein, Arabidopsis