RESUMEN
DNA methylation has a crucial role in plant development regulating gene expression and silencing of transposable elements. Maintenance DNA methylation in plants occurs at symmetrical (m)CG and (m)CHG contexts ((m) = methylated) and is maintained by DNA METHYLTRANSFERASE 1 (MET1) and CHROMOMETHYLASE (CMT) DNA methyltransferase protein families, respectively. While angiosperm genomes encode for several members of MET1 and CMT families, the moss Physcomitrella patens, serving as a model for early divergent land plants, carries a single member of each family. To determine the function of P. patens PpMET we generated ΔPpmet deletion mutant which lost (m)CG and unexpectedly (m)CCG methylation at loci tested. In order to evaluate the extent of (m)CCG methylation by MET1, we reexamined the Arabidopsis thaliana Atmet1 mutant methylome and found a similar pattern of methylation loss, suggesting that maintenance of DNA methylation by MET1 is conserved through land plant evolution. While ΔPpmet displayed no phenotypic alterations during its gametophytic phase, it failed to develop sporophytes, indicating that PpMET plays a role in gametogenesis or early sporophyte development. Expression array analysis revealed that the deletion of PpMET resulted in upregulation of two genes and multiple repetitive sequences. In parallel, expression analysis of the previously reported ΔPpcmt mutant showed that lack of PpCMT triggers overexpression of genes. This overexpression combined with loss of (m)CHG and its pleiotropic phenotype, implies that PpCMT has an essential evolutionary conserved role in the epigenetic control of gene expression. Collectively, our results suggest functional conservation of MET1 and CMT families during land plant evolution. A model describing the relationship between MET1 and CMT in CCG methylation is presented.
Asunto(s)
Bryopsida/genética , Bryopsida/metabolismo , ADN (Citosina-5-)-Metiltransferasas/metabolismo , Metilación de ADN , Proteínas de Plantas/metabolismo , Oxidorreductasas de Alcohol , Secuencia de Bases , Bryopsida/crecimiento & desarrollo , ADN (Citosina-5-)-Metiltransferasas/genética , Metilación de ADN/genética , ADN de Plantas/genética , ADN de Plantas/metabolismo , Epigénesis Genética , Regulación de la Expresión Génica de las Plantas , Genes de Plantas , Modelos Biológicos , Datos de Secuencia Molecular , Mutación , Proteínas de Plantas/genética , Plantas Modificadas GenéticamenteRESUMEN
C-5 DNA methylation is an essential mechanism controlling gene expression and developmental programs in a variety of organisms. Though the role of DNA methylation has been intensively studied in mammals and Arabidopsis, little is known about the evolution of this mechanism. The chromomethylase (CMT) methyltransferase family is unique to plants and was found to be involved in DNA methylation in Arabidopsis, maize and tobacco. The moss Physcomitrella patens, a model for early terrestrial plants, harbors a single homolog of the CMT protein family designated as PpCMT. Our phylogenetic analysis suggested that the CMT family is unique to embryophytes and its earliest known member PpCMT belongs to the CMT3 subfamily. Thus, P. patens may serve as a model to study the ancient functions of the CMT3 family. We have generated a ΔPpcmt deletion mutant which demonstrated that PpCMT is essential for P. patens protonema and gametophore development and is involved in CHG methylation as demonstrated at four distinct genomic loci. PpCMT protein accumulation pattern correlated with proliferating cells and was sub-localized to the nucleus as predicted from its function. Taken together, our results suggested that CHG DNA methylation mediated by CMT has been employed early in land plant evolution to control developmental programs during both the vegetative and reproductive haploid phases along the plant life cycle.
