Transcription generates local topological and mechanical constraints on bacterial DNA, leading to the formation of supercoiled chromosome domains. However, the global impact of transcription on chromosome organization remains unclear due to the coarse resolution of Hi-C maps (~5–10 kb). This study combines sub-kb Hi-C contact maps and chromosome engineering to visualize individual transcriptional units (TUs). The results show that TUs form discrete three-dimensional transcription-induced domains (TIDs) that impose mechanical and topological constraints on their neighboring sequences, modifying their localization and dynamics. These findings suggest that TIDs are the primary building blocks of bacterial chromosome folding, locally imposing structural and dynamic constraints. High-resolution Hi-C contact maps reveal strong heterogeneity in short-range contact signals, with bundled domains spanning 1–20 kb that correlate with transcriptional activity. These TIDs are separated by non-transcribed regions but can interact over short distances. The study also demonstrates that a single active TU can induce bundled domains and arched stripe patterns in Hi-C maps, suggesting that these structures are driven by RNA polymerase (RNAP) occupancy. Additionally, the orientation and distance between neighboring TUs affect their interaction patterns, with divergent orientations leading to self-interacting domains and convergent orientations resulting in abrupt termination of transcription tracks. Live imaging further reveals that TIDs influence the localization and mobility of flanking chromosomal regions, suggesting that transcription modulates DNA localization and imposes mechanical constraints on neighboring loci. Overall, this work highlights the importance of transcription in shaping bacterial chromosome architecture and dynamics.Transcription generates local topological and mechanical constraints on bacterial DNA, leading to the formation of supercoiled chromosome domains. However, the global impact of transcription on chromosome organization remains unclear due to the coarse resolution of Hi-C maps (~5–10 kb). This study combines sub-kb Hi-C contact maps and chromosome engineering to visualize individual transcriptional units (TUs). The results show that TUs form discrete three-dimensional transcription-induced domains (TIDs) that impose mechanical and topological constraints on their neighboring sequences, modifying their localization and dynamics. These findings suggest that TIDs are the primary building blocks of bacterial chromosome folding, locally imposing structural and dynamic constraints. High-resolution Hi-C contact maps reveal strong heterogeneity in short-range contact signals, with bundled domains spanning 1–20 kb that correlate with transcriptional activity. These TIDs are separated by non-transcribed regions but can interact over short distances. The study also demonstrates that a single active TU can induce bundled domains and arched stripe patterns in Hi-C maps, suggesting that these structures are driven by RNA polymerase (RNAP) occupancy. Additionally, the orientation and distance between neighboring TUs affect their interaction patterns, with divergent orientations leading to self-interacting domains and convergent orientations resulting in abrupt termination of transcription tracks. Live imaging further reveals that TIDs influence the localization and mobility of flanking chromosomal regions, suggesting that transcription modulates DNA localization and imposes mechanical constraints on neighboring loci. Overall, this work highlights the importance of transcription in shaping bacterial chromosome architecture and dynamics.