RESUMEN
The maize (Zea mays) RNA Polymerase IV (Pol IV) largest subunit, RNA Polymerase D1 (RPD1 or NRPD1), is required for facilitating paramutations, restricting expression patterns of genes required for normal development, and generating small interfering RNA (siRNAs). Despite this expanded role for maize Pol IV relative to Arabidopsis thaliana, neither the general characteristics of Pol IV-regulated haplotypes, nor their prevalence, are known. Here, we show that specific haplotypes of the purple plant1 locus, encoding an anthocyanin pigment regulator, acquire and retain an expanded expression domain following transmission from siRNA biogenesis mutants. This conditioned expression pattern is progressively enhanced over generations in Pol IV mutants and then remains heritable after restoration of Pol IV function. This unusual genetic behavior is associated with promoter-proximal transposon fragments but is independent of sequences required for paramutation. These results indicate that trans-generational Pol IV action defines the expression patterns of haplotypes using co-opted transposon-derived sequences as regulatory elements. Our results provide a molecular framework for the concept that induced changes to the heterochromatic component of the genome are coincident with heritable changes in gene regulation. Alterations of this Pol IV-based regulatory system can generate potentially desirable and adaptive traits for selection to act upon.
Asunto(s)
ARN Polimerasas Dirigidas por ADN/metabolismo , Epigénesis Genética , Regulación de la Expresión Génica de las Plantas , ARN de Planta/metabolismo , Zea mays/enzimología , Zea mays/genética , Alelos , Antocianinas/genética , Antocianinas/metabolismo , Ensamble y Desensamble de Cromatina , Elementos Transponibles de ADN , ARN Polimerasas Dirigidas por ADN/genética , Sitios Genéticos , Haplotipos , Patrón de Herencia , Datos de Secuencia Molecular , Fenotipo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , ARN de Planta/genética , ARN Interferente Pequeño/genética , ARN Interferente Pequeño/metabolismo , Secuencias Reguladoras de Ácidos Nucleicos , Selección GenéticaRESUMEN
Meiotically heritable epigenetic changes in gene regulation known as paramutations are facilitated by poorly understood trans-homolog interactions. Mutations affecting paramutations in maize (Zea mays) identify components required for the accumulation of 24-nucleotide RNAs. Some of these components have Arabidopsis thaliana orthologs that are part of an RNA-directed DNA methylation (RdDM) pathway. It remains unclear if small RNAs actually mediate paramutations and whether the maize-specific molecules identified to date define a mechanism distinct from RdDM. Here, we identify a novel protein required for paramutation at the maize purple plant1 locus. This required to maintain repression2 (RMR2) protein represents the founding member of a plant-specific clade of predicted proteins. We show that RMR2 is required for transcriptional repression at the Pl1-Rhoades haplotype, for accumulation of 24-nucleotide RNA species, and for maintenance of a 5-methylcytosine pattern distinct from that maintained by RNA polymerase IV. Genetic tests indicate that RMR2 is not required for paramutation occurring at the red1 locus. These results distinguish the paramutation-type mechanisms operating at specific haplotypes. The RMR2 clade of proteins provides a new entry point for understanding the diversity of epigenomic control operating in higher plants.
Asunto(s)
Proteínas de Plantas/genética , Zea mays/genética , 5-Metilcitosina/metabolismo , Arabidopsis/genética , Arabidopsis/metabolismo , Metilación de ADN/genética , Regulación de la Expresión Génica de las Plantas/genética , Haplotipos , Datos de Secuencia Molecular , Zea mays/metabolismoRESUMEN
Targeting of Saccharomyces cerevisiae Cdc24p to polarized growth sites is essential for its function. Localization of GFP-tagged Cdc24 proteins or fragments was assayed in deletion mutants of Cdc24p-interacting proteins. The boi2Delta, ent2Delta, and hua1Delta mutants showed localization defects. The tos2Delta skg6Delta double mutant displayed aberrant pre-anaphase localization to the mother-bud neck region. The same aberrant pattern was seen when potential phosphorylation sites Ser697, Thr704, and Tyr200 were mutated. The S697A mutation also resulted in phosphorylation defects in vivo. These data support roles for Boi2p, Ent2p, Hua1p, Tos2p, and for Cdc24p phosphorylation in targeting Cdc24p to growth sites.