Identification of DYRK1B as a substrate of ERK1/2 and characterisation of the kinase activity of DYRK1B mutants from cancer and metabolic syndrome

The dual-specificity tyrosine-phosphorylation- regulated kinase, DYRK1B, is expressed de novo during myogenesis, amplified or mutated in certain cancers and mutated in familial cases of metabolic syndrome. DYRK1Bis activated by cis auto-phosphorylation on tyr- osine-273 (Y273) within the activation loop during translation but few other DYRK1Bphosphorylation sites have been characterised to date. Here, we demonstrate that DYRK1Balso undergoes trans- autophosphorylationon serine-421 (S421) in vitro and in cells and that this site contributes to DYRK1Bkinase activity. Whilst a DYRK1BS421A mutant was completely defective for p-S421 in cells, DYRK1Binhibitors caused only a partial loss of p-S421 suggesting the existence of an additional kinase that could also phosphorylate DYRK1BS421. Indeed, a catalytically inactive DYRK1BD239A mutant exhibited very low levels of p-S421 in cells but this was increased by KRAS G12V. In addition, selective activation of the RAF-MEK1/2-ERK1/2 signalling pathway rapidly increased p-S421 in cells whereas activation of the stress kinases JNK or p38 could not. S421 resides within a Ser- Pro phosphoacceptor motif that is typical for ERK1/2 and recombinant ERK2 phosphorylated DYRK1Bat S421 in vitro. Our results show that DYRK1Bis a novel ERK2 substrate, uncovering new links between two kinases involved in cell fate decisions. Finally, we show that DYRK1Bmutants that have recently been described in cancer and metabolic syndrome exhibit normal or reduced intrinsic kinase activity.