Engineered Covalent Inactivation of TFIIH-Kinase Reveals an Elongation Checkpoint and Results in Widespread mRNA Stabilization

During transcription initiation, the TFIIH-kinase Kin28/Cdk7 marks RNA polymerase II (Pol II) by phosphorylating the C-terminal domain (CTD) of its largest subunit. Here we describe a structure-guided chemical approach to covalently and specifically inactivate Kin28 kinase activity in vivo. This method of irreversible inactivation recapitulates both the lethal phenotype and the key molecular signatures that result from genetically disrupting Kin28 function in vivo. Inactivating Kin28 impacts promoter release to differing degrees and reveals a “checkpoint” during the transition to productive elongation. While promoter-proximal pausing is not observed in budding yeast, inhibition of Kin28 attenuates elongation-licensing signals, resulting in Pol II accumulation at the +2 nucleosome and reduced transition to productive elongation. Furthermore, upon inhibition, global stabilization of mRNA masks different degrees of reduction in nascent transcription. This study resolves long-standing controversies on the role of Kin28 in transcription and provides a rational approach to irreversibly inhibit other kinases in vivo. [Display omitted] •A general approach for covalent chemical inhibition of kinases in vivo•Targeted inhibition of Kin28 reveals an elusive elongation checkpoint in yeast•Varying impact on promoter escape and transition to productive elongation•Stabilization of existing mRNA buffers/masks reduction in nascent transcripts Rodriguez-Molina et al. describe a general strategy for irreversible inhibition of kinases in vivo. Inhibition of Kin28/CDK7 reduces nascent transcription, increases stability of existing mRNA, and reveals an underappreciated role for Kin28 in priming Pol II for productive transcription elongation.