Experimental demonstration of operon formation catalyzed by insertion sequence

Operons are a hallmark of the genomic and regulatory architecture of prokaryotes. However, the mechanism by which two genes placed far apart gradually come close and form operons remains to be elucidated. Here, we propose a new model of the origin of operons: Mobile genetic elements called insertion sequences can facilitate the formation of operons by consecutive insertion–deletion–excision reactions. This mechanism barely leaves traces of insertion sequences and thus difficult to detect in nature. In this study, as a proof-of-concept, we reproducibly demonstrated operon formation in the laboratory. The insertion sequence IS3 and the insertion sequence excision enhancer are genes found in a broad range of bacterial species. We introduced these genes into insertion sequence-less Escherichia coli and found that, supporting our hypothesis, the activity of the two genes altered the expression of genes surrounding IS3, closed a 2.7 kb gap between a pair of genes, and formed new operons. This study shows how insertion sequences can facilitate the rapid formation of operons through locally increasing the structural mutation rates and highlights how coevolution with mobile elements may shape the organization of prokaryotic genomes and gene regulation.