Skip to main page content
U.S. flag

An official website of the United States government

Dot gov

The .gov means it’s official.
Federal government websites often end in .gov or .mil. Before sharing sensitive information, make sure you’re on a federal government site.

Https

The site is secure.
The https:// ensures that you are connecting to the official website and that any information you provide is encrypted and transmitted securely.

Access keys NCBI Homepage MyNCBI Homepage Main Content Main Navigation
. 2001 Aug;13(8):1919-28.
doi: 10.1105/tpc.010064.

Maize chromomethylase Zea methyltransferase2 is required for CpNpG methylation

C M Papa  1 , N M Springer, M G Muszynski, R Meeley, S M Kaeppler
Affiliations

Maize chromomethylase Zea methyltransferase2 is required for CpNpG methylation

C M Papa et al. Plant Cell. 2001 Aug.

Abstract

A cytosine DNA methyltransferase containing a chromodomain, Zea methyltransferase2 (Zmet2), was cloned from maize. The sequence of ZMET2 is similar to that of the Arabidopsis chromomethylases CMT1 and CMT3, with C-terminal motifs characteristic of eukaryotic and prokaryotic DNA methyltransferases. We used a reverse genetics approach to determine the function of the Zmet2 gene. Plants homozygous for a Mutator transposable element insertion into motif IX had a 13% reduction in methylated cytosines. DNA gel blot analysis of these plants with methylation-sensitive restriction enzymes and bisulfite sequencing of a 180-bp knob sequence showed reduced methylation only at CpNpG sites. No reductions in methylation were observed at CpG or asymmetric sites in heterozygous or homozygous mutant plants. Our research shows that chromomethylase Zmet2 is required for in vivo methylation of CpNpG sequences.

PubMed Disclaimer

Figures

Figure 1.
Figure 1.
Maize Zmet2 Encodes a Chromomethylase. (A) The conserved methyltransferase motifs of the ZMET2 and ZMET5 inferred amino acid sequences are aligned with the Arabidopsis chromomethylases CMT1, CMT2, and CMT3. Black shading indicates identical residues, and gray shading indicates similarity. Dashes in the sequences represent gaps introduced by CLUSTAL W to optimize the alignments. Alignments were processed by BOXSHADE. The locations of the six conserved methylase motifs are indicated above the sequences. The chromodomain is located upstream of and adjacent to motif IV. The location of the Mu insertion in the zmet2-m1::Mu allele is indicated by an arrowhead above the sequences. (B) Relationships of maize and Arabidopsis chromomethylases. The aligned conserved methyltransferase motifs of ZMET2, ZMET5, CMT1, CMT2, and CMT3 (from the beginning of motif I to the end of motif X) were analyzed using PHYLIP. The resulting tree is shown with bootstrap values indicated at the nodes. (C) Diagrams of methyltransferase proteins. The chromomethylases of maize and Arabidopsis are shown with Zmet1 and Zmet3. The location of the chromodomains (CD), the conserved methyltransferase motifs (I, IV, and VI to X), the BAH domains, and the ubiquitin-associated domains (UBA) are indicated by different shading.
Figure 2.
Figure 2.
Gel Blot Analysis of Repetitive DNA Methylation Patterns. Decreased methylation is observed in mutant plants (− −) relative to nonmutant plants (+ +) digested with MspI, which is sensitive to methylation at mCpCpG sequences. No changes in methylation patterns at mCpG sites are observed in mutant plants, as indicated by the lack of digestion with HpaII. Plants heterozygous for zmet2-m1::Mu (+ −) also show decreases at mCpCpG sites. DNA gel blots were hybridized with probes for repetitive DNA: the 9-kb 26s-5.8s-17s ribosomal repeat (A), the 5s ribosomal repeat (B), and the centromeric repeat pSau3A9 (C).
Figure 3.
Figure 3.
The DNA Methylation Patterns of Both Strands of the 180-bp Knob Repeat Were Determined by Direct Genomic Bisulfite Sequencing. CpG, CpNpG, and asymmetric methylation all were detected in the knob sequence of plants wild type for Zmet2. CpG dinucleotides are indicated by ovals, and CpNpG trinucleotides are indicated by rectangles. The symbols above and below the alignment indicate the amount of DNA methylation observed in wild-type and homozygous zmet2-m1::Mu plants. When only one symbol is shown, the methylation was the same in wild-type and zmet2-m1::Mu plants. When the methylation status differed, two symbols are shown, with the top symbol showing the methylation status of that base in zmet2-m1::Mu plants and the bottom symbol showing the methylation status of that base in wild-type siblings. The only differences in methylation were found at CpNpG sequences. No changes in CpG or asymmetric methylation were observed.
See this image and copyright information in PMC

Similar articles

See all similar articles

Cited by

See all "Cited by" articles

References

    1. Akhtar, A., Zink, D., and Becker, P.B. (2000). Chromodomains are protein–RNA interaction modules. Nature 407, 405–409. - PubMed
    1. Banks J.A., Masson, P., and Fedoroff, N. (1988). Molecular mechanisms in the developmental regulation of the Suppressor-mutator transposable element. Genes Dev. 2, 1364–1380. - PubMed
    1. Bensen, R.J., Johal, G.S., Crane, V.C., Tossberg, J.T., Schnable, P.S., Meeley, R.B., and Briggs, S.B. (1995). Cloning and characterization of the maize An1 gene. Plant Cell 7, 75–84. - PMC - PubMed
    1. Callebaut, I., Courvalin, J.C., and Mornon, J.P. (1999). The BAH (bromo-adjacent homology) domain: A link between DNA methylation, replication and transcriptional regulation. FEBS Lett. 446, 189–193. - PubMed
    1. Cao, X., Springer, N.S., Muszynski, M.G., Phillips, R.L., Kaeppler, S., and Jacobsen, S.E. (2000). Conserved plant genes with similarity to mammalian de novo DNA methyltransferases. Proc. Natl. Acad. Sci. USA 97, 4979–4984. - PMC - PubMed

Publication types

Associated data