Disruption of their palindromic arrangement leads to selective loss of DNAmethylation in inversely repeated gus transgenes in Arabidopsis

The transgene locus KH15, which is highly susceptible to silencing in Arabi dopsis thaliana, contains two inversely repeated beta -glucuronidase (gus) genes separated by a palindromic sequence and has a low GUS activity, was f ound to be heavily methylated in the gus coding sequence and in the center of the inverted repeat. The locus KHsb67, which is less prone to silencing, was found to be less densely methylated in the non-repetitive region that separates the inversely repeated gus genes. After the removal of one of the gus genes by Cre-mediated recombination, methylation in both loci decrease d or was totally lost. Despite the presence of a 732-bp palindromic sequenc e in the deletion line derived from KH15, this sequence was not methylated. Whereas the KH15 locus triggers methylation of homologous gus genes when p laced in trans to them, the deletion derivative did not. suggesting that th e capacity for cross-talk was severely affected by disruption of the palind romic arrangement. This result suggests that the transcribed palindromic se quences are required to maintain the methylation of both symmetrically and non-symmetrically arranged cytosines.