Post-transcriptional gene silencing of chalcone synthase in transgenic petunias, cytosine methylation and epigenetic variation

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Post-transcriptional degradation of chalcone synthase RNA transcripts often occurs in petunia plants with additional copies of chalcone synthase genes. The cytosine methylation status of selected restriction endonuclease sites was studied in the chalcone synthase endogenous genes and transgenes of these plants. Methylation patterns varied between transformants and within a series of epigenetically related plants. There was heterogeneity in promoter transgene methylation patterns within and/or between cells, between leaves and flowers and between tandemly duplicated sequences. Thus methylation patterns are determined by more than the local DNA sequence or site of insertion. A site in the 5′ enhancer of the 35S promoter was methylated more frequently in white flowers than in purple flowers. An EcoRII site within the coding sequence of the endogenous ChsA genes is developmentally regulated, being frequently methylated in leaves but rarely in petals of wild-type petunias or stable purple flowered transgenic plants. However, in petals of many transgenic plants, especially those that show extensive post transcriptional RNA degradation, the site is frequently methylated. Thus the transgenes influence developmental methylation of the endogenous genes. In the transgenes the EcoRII site is usually more frequently methylated in petals than in leaves. RNA degradation in petals is correlated in one set of epigenetically related transgenic plants with hypomethylation at the transgene EcoRII site in leaves. Possible relationships between the cytosine methylation patterns in vegetative and floral cells, epigenetic states of ChsA genes and the control of RNA transcript degradation are discussed.

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