Comparison of REST Cistromes across Human Cell Types Reveals Common and Context-Specific Functions

Abstract

Recent studies have shown that the transcriptional functions of REST are much broader than repressing neuronal genes in non-neuronal systems. Whether REST occupies similar chromatin regions in different cell types and how it interacts with other transcriptional regulators to execute its functions in a context-dependent manner has not been adequately investigated. We have applied ChIP-seq analysis to identify the REST cistrome in human CD4+ T cells and compared it with published data from 15 other cell types. We found that REST cistromes were distinct among cell types, with REST binding to several tumor suppressors specifically in cancer cells, whereas 7% of the REST peaks in non-neuronal cells were ubiquitously called and <25% were identified for ≥5 cell types. Nevertheless, using a quantitative metric directly comparing raw ChIP-seq signals, we found the majority (∼80%) was shared by ≥2 cell types. Integration with RNA-seq data showed that REST binding was generally correlated with low gene expression. Close examination revealed that multiple contexts were correlated with reduced expression of REST targets, e.g., the presence of a cognate RE1 motif and cellular specificity of REST binding. These contexts were shown to play a role in differential corepressor recruitment. Furthermore, transcriptional outcome was highly influenced by REST cofactors, e.g., SIN3 and EZH2 co-occupancy marked higher and lower expression of REST targets, respectively. Unexpectedly, the REST cistrome in differentiated neurons exhibited unique features not observed in non-neuronal cells, e.g., the lack of RE1 motifs and an association with active gene expression. Finally, our analysis demonstrated how REST could differentially regulate a transcription network constituted of miRNAs, REST complex and neuronal factors. Overall, our findings of contexts playing critical roles in REST occupancy and regulatory outcome provide insights into the molecular interactions underlying REST's diverse functions, and point to novel roles of REST in differentiated neurons.

Author Summary

The RE-1 silencing transcription factor (REST) binds to DNA and has been shown to repress neuronal genes in non-neuronal systems, but more recent studies have expanded its functions much beyond this. At the molecular level, REST acts cooperatively with other proteins to execute its transcriptional regulatory roles. The dynamics of REST binding and cofactor recruitment and its association with the underlying DNA sequence remain unclear. Here, we have applied chromatin immunoprecipitation and deep sequencing to identify REST binding across 16 different cell types, including neurons. Our results demonstrate that REST binding events are dynamic and quite distinct among cells and that REST binding is generally associated with low gene expression. Closer examination finds that the context of the DNA sequence at REST bound sites is associated with the lower expression of REST-associated targets and that different contexts correlate with different cofactor recruitment. These in turn have an effect on the expression of REST targets. REST targets in human neurons, however, are drastically different from those in other cell types. These findings provide insights into the effect of genomic and cellular contexts on REST's diverse functions and point to distinct and novel roles for REST in neurons.