Changes in T-DNA methylation and expression are associated with phenotypic variation and plant regeneration in a crown gall tumor line

Abstract

Phenotypic variation of an octopine-type crown gall tumor line resulting from changes in the pattern of T-DNA methylation and expression is described. Variants that grow as unorganized callus always express T-DNA transcripts 1 and 2. In shoot-forming variants (teratomas) only T-DNA transcript 4 is expressed. This line also regenerates normal-appearing, rooted plants in which all T-DNA expression is suppressed. Tissues from these plants require phytohormones for growth in vitro. These plants are self-fertile and transmit T-DNA through meiosis, and T-DNA suppression is maintained in the next generation. After treatment of regenerated plant tissue with 5-azacytidine, an inhibitor of DNA methylation, T-DNA transcription and phytohormone-independent tumorous growth resume. The T-DNA of cell lines in which T-DNA is not expressed is highly methylated, whereas the level of T-DNA methylation is reduced in 5-azacytidine treated cells that resume T-DNA expression and phytohormone-independent growth. The correlation between the degree of T-DNA methylation and the level of T-DNA expression indicates that hypermethylation is responsible for the suppression of T-DNA transcription.