Computational and Functional Analysis of Structural Features in the ZAK alpha Kinase

The kinase ZAKa acts as the proximal sensor of translational impairment and ribotoxic stress, which results in the activation of the MAP kinases p38 and JNK. Despite recent insights into the functions and binding partners of individual protein domains in ZAKa, the mechanisms by which ZAKa binds ribosomes and becomes activated have remained elusive. Here, we highlight a short, thrice-repeated, and positively charged peptide motif as critical for the ribotoxic stress-sensing function of the Sensor (S) domain of ZAKa. We use this insight to demonstrate that the mutation of the SAM domain uncouples ZAKa activity from ribosome binding. Finally, we use 3D structural comparison to identify and functionally characterize an additional folded domain in ZAKa with structural homology to YEATS domains. These insights allow us to formulate a model for ribosome-templated ZAKa activation based on the re-organization of interactions between modular protein domains. In sum, our work both advances our understanding of the protein domains and 3D architecture of the ZAKa kinase and furthers our understanding of how the ribotoxic stress response is activated.