Examining the Roles of Genomic Context and Endogenous Regulatory Elements on IS1 Transposition Within the Escherichia...

Insertion sequence (IS) elements are key drivers of bacterial genome plasticity, yet the overall regulation of their transposition remains poorly understood. This is especially true for the multiple-layer regulation at the donor site, which has been largely overlooked. Using multiple mutation assays, genetic manipulations and reporter genes, this study focuses on characterizing how endogenous DNA sequences, transcriptional and translational factors, and genomic context regulate IS1 transposition from its donor site. Out of six elements within the chromosome of E. coli strain BW25113, IS1A and IS1E (both with the consensus sequence) contribute to over 99.9% of the overall IS1 transposition within the genome while the other four elements without the non-consensus sequence are essentially incapable of transposing. Inducing a ribosomal -1 frameshift at the A₆C motif increases transposition over 1000-fold, but this enhancement is largely reversed by restoring InsA-mediated transcriptional regulation. Strikingly, genomic sequences flanking IS1 elements appreciably modulate transposition by promoting transcription or facilitating formation of transpososomes, a phenomenon that remains under-studied. Finally, IS1 was confirmed to undergo replicative transposition intramolecularly, a mechanism shown here to be independent of transposase levels in the cell. These findings contribute to our understanding of mobile genetic element regulation and potentially offer strategies for mitigating their potentially harmful effects.