Abstract
Homeodomain-interacting protein kinases (HIPKs) belong to the CMGC kinase family and are closely related to dual-specificity tyrosine phosphorylation-regulated kinases (DYRKs). HIPKs are regulators of various signaling pathways and involved in the pathology of cancer, chronic fibrosis, diabetes, and multiple neurodegenerative diseases. Here, we report the crystal structure of HIPK3 in its apo form at 2.5 Å resolution. Recombinant HIPKs and DYRK1A are auto-activated and phosphorylate the negative elongation factor SPT5, the transcription factor c-Myc, and the C-terminal domain of RNA polymerase II, suggesting a direct function in transcriptional regulation. Based on a database search, we identified abemaciclib, an FDA-approved Cdk4/Cdk6 inhibitor used for the treatment of metastatic breast cancer, as potent inhibitor of HIPK2, HIPK3, and DYRK1A. We determined the crystal structures of HIPK3 and DYRK1A bound to abemaciclib, showing a similar binding mode to the hinge region of the kinase as observed for Cdk6. Remarkably, DYRK1A is inhibited by abemaciclib to the same extent as Cdk4/Cdk6 in vitro, raising the question of whether targeting of DYRK1A contributes to the transcriptional inhibition and therapeutic activity of abemaciclib.
Highlights
Homeodomain-interacting protein kinases (HIPKs) belong to the CMGC kinase family and are closely related to dual-specificity tyrosine phosphorylation-regulated kinases (DYRKs)
To determine the structure of the human HIPK3 kinase domain, residues 159–562 were expressed in baculovirus infected Spodoptera frugiperda 9 (Sf9) insect cells
Despite its presence in the crystallization condition, no crystallographic density was seen for ADP or magnesium, implying that the kinase is in the nucleotide free apo-state
Summary
Homeodomain-interacting protein kinases (HIPKs) belong to the CMGC kinase family and are closely related to dual-specificity tyrosine phosphorylation-regulated kinases (DYRKs). HIPKs belong to the CMGC group of serine/threonine kinases and are part of the dualspecificity tyrosine phosphorylation-regulated kinase (DYRK). DYRKs are named after their characteristic dual-specificity, as they auto-phosphorylate a conserved tyrosine in their activation loop, but phosphorylate substrates on serine and threonine residues[2]. For the class II DYRK ortholog from D. melanogaster it was demonstrated that the critical tyrosine is cis-auto-phosphorylated by the nascent kinase in a transitory intermediate state during maturation at the ribosome[3]. The phosphorylation of the critical tyrosine was later confirmed for members of all DYRK subfamilies by biochemical and structural analyses[4,5,6,7]. Within the HIPK family, HIPK1 and HIPK2 are the most closely related members, sharing about 93%
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