Molecular Mechanisms of DKRK and GSK3 Autophosphorlation
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DYRK autophosphorlation - During maturation of the DYRK molecule, prior to release from the ribosome, the nascent kinase forms a transitional intermediate. The intermediate functions exclusively as an intra-molecular tyrosine kinase and phosphorylates one substrate only, an essential tyrosine residue in the molecule’s own activation loop. The fully translated, activation loop-phosphorylated DYRK then assumes its mature conformation. The intra-molecular tyrosine kinase activity is lost, and the kinase functions exclusively as an inter-molecular serine/threonine kinase. The different residue and substrate specificities and sensitivities to small molecule inhibitors of the intermediate and mature kinase are outlined.
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GSK3 autophosphorlation - GSK3 is translated and released from the ribosome. Shortly thereafter, the protein forms a transitional intermediate that exhibits canonical tyrosine kinase activity toward the molecules own activation loop Tyr216. Phosphorylation of Tyr216 is intra-molecular and requires the chaperone activity of hsp90. After protein maturation and Tyr216 phosphorylation, mature GSK3 then functions as a serine/threonine kinase that phosphorylates substrates via an inter-molecular mechanism. The residue phosphorylated, its sequence context, and sensitivities to inhibitors of the intermediate and mature kinase are shown.
Our lab is interested in the mechanism by which dual-specificity tyrosine-phosphorylation-regulated protein kinases (DYRKs), and glycogen synthase kinase 3s (GSK3s) are regulated.
The phosphorylation of proteins on serine, threonine or tyrosine residues is arguably the most pervasive and important post-translational modification affecting cell life. Phosphorylation plays key roles in diverse processes such as cellular proliferation, differentiation, motility, metabolism, and gene expression. Abnormal phosphorylation contributes to a very large number of human diseases, including cancer, developmental abnormality, diabetes, inflammation, and neurological disorders. The DYRK family of kinases for example, is strongly implicated in Down Syndrome, microcephaly, cancer, and Alzheimer's disease.
For many diseases, protein kinases represent a point for therapeutic intervention and ~30% of all research spending on drug development focuses on protein kinase inhibitors. GSK3 is implicated in a variety of human diseases and many pharmaceutical companies are striving to develop small molecule inhibitors of this protein for the treatment of diabetes, Alzheimers, bipolar affective disorders and other neurological diseases. Understanding the molecular basis of kinase regulation therefore represents a fundamental challenge for basic and applied science. Our lab uses molecular biology and Drosophila genetics to better understand fundamental mechanisms of protein kinase regulation and the role these enzymes play in developmental signal transduction pathways.
One of the most important modes of protein kinase regulation is the phosphorylation of key residues in the activation loop of the kinase domain; an event that can increase kinase activity by 2-5 orders of magnitude. Previous work in our laboratory identified a novel mechanism whereby DYRKs and GSK3 enzymes are activated. DYRKs and GSK3 employ a short-lived transitional intermediate form of the kinase to autophosphorylate an essential tyrosine in the activation loop using an intra-molecular mechanism. This intra-molecular event occurs during the initial folding of the protein converting the kinases into specific inter-molecular serine/threonine kinases. The intermediates differ in residue and substrate specificity, and sensitivity to small-molecule inhibitors, compared to their mature counterpart. The identification of protein kinase intermediates demonstrates that DYRKs, GSK3, and other kinases activated in this way can be targeted at two levels; the transitory intermediate and the mature active enzyme.
Publications
Lochhead, P.A., G. Sibbet, N. Morrice and V. Cleghon. 2005.
Activation-loop autophosphorylation is mediated by a novel transitional intermediate form of DYRKs. Cell 121:925-36.
Lochhead, P.A., R. Kinstrie, G. Sibbet, T. Rawjee, N. Morrice and V. Cleghon, 2006.
GSK3β activation-loop autophosphorylation is mediated by a chaperone-dependent transitional intermediate. Molecular Cell 24:627-633.
Kinstrie, R., P.A. Lochhead, G. Sibbet, N. Morrice and V. Cleghon. 2006.
dDYRK2 and Minibrain interact with the chromatin remodelling factors SNR1 and TRX. Biochem J, 2006. 398:45-54.
Contact
Dr. Vaughn Cleghon
Cincinnati Children’s Hospital Medical Center
Division of Developmental Biology
3333 Burnet Ave, MLC 7007
Cincinnati, OH 45229
Phone 513-803-0470
Fax 513-636-4317
E-Mail vaughn.cleghon@cchmc.org
