Fragile X (CGG)n repeats induce a transcriptional repression in cis upon a linked promoter:: Evidence for a chromatin mediated effect -: art. no. 3

Background: Expansion of an unstable (CGG)(n) repeat to over 200 triplets within the promoter region of the human FMR1 gene leads to extensive local methylation and transcription silencing, resulting in the loss of FMRP protein and the development of the clinical features of fragile X syndrome. The causative link between (CGG)(n) expansion, methylation and gene silencing is unknown, although gene silencing is associated with extensive changes to local chromatin architecture. Results: In order to determine the direct effects of increased repeat length on gene transcription in a chromatin context, we have examined the influence of FMR1 ( CGG) n repeats upon transcription from the HSV thymidine kinase promoter in the Xenopus laevis oocyte. We observe a reduction in mRNA production directly associated with increasing repeat length, with a 90% reduction in mRNA production from arrays over 100 repeats in length. Using a kinetic approach, we show that this transcriptional repression is concomitant with chromatin maturation and, using in vitro transcription, we show that chromatin formation is a fundamental part of the repressive pathway mediated by (CGG)(n) repeats. Using Trichostatin A, a histone deacetylase inhibitor, we show reactivation of the silenced promoter. Conclusions: Thus, isolated fragile X associated (CGG)(n) repeat arrays can exert a modifying and transcriptionally repressive influence over adjacent promoters and this repressive phenomenon is, in part, mediated by histone deacetylation.