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.