MTBP allosterically activates Cdk8-CycC kinase activity

How the approximately 300 human protein kinases identify their dedicated substrates despite the significant overlap of their phosphorylation consensus sequences is relevant for nearly all cellular processes. We show here that the Cdk8/19-CycC kinase uses mutually exclusive targeting and activation factors to facilitate distinct cellular roles. The Med12 protein is known to target Cdk8/19-CycC to the mediator of transcription complex to control the transcription of specific gene sets upon respective stimuli. We describe that a second Cdk8/19-CycC targeting factor exists, the replication origin firing regulator MTBP that targets the kinase to Med12-independent cellular roles. Both Med12 and MTBP constitute allosteric activators of the enzymatic Cdk8/19-CycC kinase activity in vitro. We describe the structural basis of this activation that involves distinct mechanisms how Med12 and MTBP reposition the T-loop of the kinase independently of T-loop phosphorylation – the canonical mechanism of CDK kinase activation. Our results support the following model: the Cdk8/19-CycC dimer alone has low enzymatic activity, which may help avoid off-target phosphorylation. Med12, MTBP and potentially other as yet unidentified accessory factors, target the kinase to distinct molecular environments, at the same time activating kinase activity for efficient substrate phosphorylation. Our work establishes an unusual mechanism of CDK kinase control and change the current paradigm how Cdk8/19-CycC selects its substrates. Substrate selection of the kinase may be relevant for cancer biology and therapy because Cdk8 is a well-established colorectal cancer promoting factor.