Transcription-factor-dependent enhancer transcription defines a gene regulatory network for cardiac rhythm

Abstract

The noncoding genome is pervasively transcribed. Noncoding RNAs (ncRNAs) generated from enhancers have been proposed as a general facet of enhancer function and some have been shown to be required for enhancer activity. Here we examine the transcription-factor-(TF)-dependence of ncRNA expression to define enhancers and enhancer-associated ncRNAs that are involved in a TF-dependent regulatory network. TBX5, a cardiac TF, regulates a network of cardiac channel genes to maintain cardiac rhythm. We deep sequenced wildtype and <i>Tbx5</i>-mutant mouse atria, identifying ~2600 novel <i>Tbx5</i>-dependent ncRNAs. Tbx5-dependent ncRNAs were enriched for tissue-specific marks of active enhancers genome-wide. Tbx5-dependent ncRNAs emanated from regions that are enriched for TBX5-binding and that demonstrated Tbx5-dependent enhancer activity. <i>Tbx5</i>-dependent ncRNA transcription provided a quantitative metric of <i>Tbx5</i>-dependent enhancer activity, correlating with target gene expression. We identified <i>RACER</i>, a novel <i>Tbx5</i>-dependent long noncoding RNA (lncRNA) required for the expression of the calcium-handling gene <i>Ryr2</i>. We illustrate that TF-dependent enhancer transcription can illuminate components of TF-dependent gene regulatory networks.