Abstract
Transcription commonly occurs in bursts, with alternating productive (ON) and quiescent (OFF) periods determining mRNA production rates. However, how bursting dynamics regulate transcription is not well understood. Here, we conduct real-time measurements of endogenous transcriptional bursting with single-mRNA sensitivity. Using the diverse transcriptional activities present in early Drosophila embryos, we find stringent relationships between bursting parameters. Specifically, ON and OFF durations are tightly coupled, and each level of gene activity is associated with a characteristic combination of these periods. Lowly transcribing alleles primarily adjust OFF periods (burst frequency), while highly transcribing alleles tune ON periods (burst size). These relationships persist across developmental stages, body-axis positions, cis-regulatory or trans-regulatory perturbations and bursting dynamics observed in other species. Our findings suggest a mechanistic constraint that governs bursting dynamics, challenging the view that regulatory processes independently control distinct parameters.