Abstract
Transcriptional silencing selectively impedes gene expression. Silencing is often accompanied by replication delay and can be prevented by replicator sequences. Here we report a replicator-binding protein complex involved in the prevention of transcriptional silencing. The protein complex interacts with an essential asymmetric region within the human β-globin Rep-P replicator and includes hnRNP C1/C2, SWI/SNF complex, and MeCP1, which are members of the locus control region (LCR)-associated remodeling complex (LARC). Interaction between LARC and Rep-P prevented transcriptional silencing and replication delay. Transgenes that did not contain the asymmetric LARC-binding region of Rep-P replicated late and exhibited stable silencing that could not be affected by a DNA methylation inhibitor. In contrast, transgenes that contain a mutation of the asymmetric region of Rep-P that could not bind LARC exhibited a silent state that could transiently be reactivated by DNA demethylation. The effect of DNA demethylation was transient, and prolonged exposure to a methylation inhibitor induced distinct, stable, methylation-independent silencing. These observations suggest that the interaction of LARC complex with replicators plays a role in preventing gene silencing and provides support for a novel, epigenetic mechanism of resistance to methylation inhibitors.