Abstract
Repetitive extragenic palindromes (REPs) are the most abundant repetitive noncoding elements in the E. coli genome. Despite their abundance, the primary function of REPs has remained unclear. At different times, REPs have been proposed to contribute to chromosome organization, mRNA decay regulation, and transcription termination, among other functions. Here, we show that the model REP, REP325, does not measurably compact the chromosome but instead acts as a 3'UTR-associated transcription regulator within the yjdMN operon, functioning both as a partially Rho-dependent terminator that limits transcription into the downstream yjdN gene and as an mRNA stabilizer that protects the upstream yjdM transcript from degradation. This dual role in controlling both transcriptional readthrough and susceptibility to decay provides a framework that reconciles several previously conflicting observations about REP function. Our genome-wide RNA-seq analysis further reveals that REPs with more canonical sequence and hairpin structures are more often associated with upstream-biased expression in tandem gene pairs, and that REPs positioned between convergent genes correlate with elevated expression of both genes. The large variance in expression patterns in both gene pair configurations is consistent with context-dependent termination and degradation blocking. Similarly, REPs do not uniformly affect mRNA half-lives. Because REP locations vary between E. coli strains, REPs likely contribute to regulatory diversity by tuning gene expression without altering protein-coding sequences or promoter regions, opening new avenues for modulating gene expression through REP-mediated transcription regulation.