Integrator loss leads to dsRNA formation that triggers the integrated stress response.

Integrator (INT) is a metazoan-specific complex that targets promoter-proximally paused RNA polymerase II (RNAPII) for termination, preventing immature RNAPII from entering gene bodies and functionally attenuating transcription of stress-responsive genes. Mutations in INT subunits are associated with many human diseases, including cancer, ciliopathies, and neurodevelopmental disorders, but how reduced INT activity contributes to disease is unknown. Here, we demonstrate that the loss of INT-mediated termination in human cells triggers the integrated stress response (ISR). INT depletion causes upregulation of short genes such as the ISR transcription factor activating transcription factor 3 (ATF3). Further, immature RNAPII that escapes into genes upon INT depletion is prone to premature termination, generating incomplete pre-mRNAs with retained introns. Retroelements within retained introns form double-stranded RNA (dsRNA) that is recognized by protein kinase R (PKR), which drives ATF4 activation and prolonged ISR. Critically, patient cells with INT mutations exhibit dsRNA accumulation and ISR activation, thereby implicating chronic ISR in diseases caused by INT deficiency.