The metazoan-specific acetyltransferase p300/CBP is involved in activating signal-induced, enhancer-mediated transcription of cell-type-specific genes. However, the global kinetics and mechanisms of p300/CBP activity-dependent transcription activation remain poorly understood. We performed genome-wide, time-resolved analyses to show that enhancers and super-enhancers are dynamically activated through p300/CBP-catalyzed acetylation, deactivated by the opposing deacetylase activity, and kinetic acetylation directly contributes to maintaining cell identity at very rapid (minutes) timescales. The acetyltransferase activity is dispensable for the recruitment of p300/CBP and transcription factors but essential for promoting the recruitment of TFIID and RNAPII at virtually all enhancers and enhancer-regulated genes. This identifies pre-initiation complex assembly as a dynamically controlled step in the transcription cycle and reveals p300/CBP-catalyzed acetylation as the signal that specifically promotes transcription initiation at enhancer-regulated genes. We propose that p300/CBP activity uses a “recruit-and-release” mechanism to simultaneously promote RNAPII recruitment and pause release and thereby enables kinetic activation of enhancer-mediated transcription. Display omitted •p300/CBP and deacetylase activities regulate dynamic (de)activation of enhancers•p300/CBP-catalyzed acetylation promotes PIC assembly and RNAPII recruitment•BRD4 acts as a p300/CBP downstream effector to promote RNAPII pause release•Coupling of RNAPII recruitment and pause release enables rapid enhancer activation A systems-wide analysis reveals that enhancers are activated by p300/CBP-catalyzed acetylation and deactivated by deacetylation. p300/CBP activity promotes pre-initiation complex assembly and RNAPII recruitment independently of its previously known function in BRD4-dependent pause release. By simultaneously promoting transcription initiation and elongation, p300/CBP activity drives the rapid activation of enhancers and super-enhancers.