Characterization of carrot nuclear proteins that exhibit specific binding affinity towards conventional and non-conventional DNA methylation

DNA methylation is associated with transcriptional silencing in vertebrates and plants. In mammals, the effects of methylation are mediated by a family of methyl-CpG-binding proteins. In plants the mechanisms by which methylation represses transcription are still not clear. In this paper we describe protein factors in carrot nuclear extracts exhibiting specific affinities for conventional or non-conventional methylation acceptor sites. We characterized two classes of proteins: The first, dcMBP1 (Daucus carota methylated DNA-binding protein 1), shows high affinity for sequences containing 5-methylcytosine; the second, dcMBP2 (Daucus carota methylated DNA-binding protein 2), efficiently complexes sequences containing 5-methylcytosine in both CpXpX and CpXpG trinucleotides and shows much lower affinity for 5-methyl CpG dinucleotides. Both dcMBP1 and dcMBP2 are abundant proteins differing in molecular weight and binding features. Their activities are modulated during carrot vegetative cell growth and somatic embryo development. This is the first time that, in either plants or mammals, proteins exhibiting specific binding affinities for conventional or non-conventional DNA methylation have been shown. Based on these results, the possibility that both the extent and the context of the methylation might contribute to modulate gene expression is discussed.