Doctoral dissertation

Juuli Raivola: Molecular Regulation of Janus Kinases (JAKs) : Focus on the Pseudokinase Domain

Juuli Raivola
The Janus kinase-signal transducer and activator of transcription (JAK-STAT) pathway mediates the transduction of over 50 cytokines that regulate cell survival, proliferation, migration, gene expression and other vital processes such as immune response. On the other hand, defects in the JAK-STAT signalling have severe impacts. Activating JAK (JAK1-3 and tyrosine kinase 2, TYK2) mutations cause haematological cancers and myeloproliferative disorders while impaired JAK- signalling leads to severe combined immunodeficiency (SCID) and autoimmune diseases.

The results presented in the thesis of Juuli Raivola focus on the JAK pseudokinase domain (JH2) that is inactive but has a crucial role in regulating the JAK activity. Our biochemical and cell-based studies show that all JAK JH2s bind ATP, but that the binding properties vary among the JAK-family. Clinical and structure-based mutation studies show that modulation of JH2 can be used to effectively alter the activity. In addition, by introducing homologous mutations into all JAK members, we observed that an individual JAK can have varying functions depending of the signaling systems it is attached to. The results highlight that each JAK within the signaling complex, and specifically JH2, is important for the signaling.

As no well-defined structures exist of the full-length JAKs, a molecular dynamic (MD) simulation model of the full-length JAK2 with erythropoietin receptor was constructed in collaboration with D.E. Shaw research (N.Y.). The model depicts JAK2 in different states of activation and is supported by structure-based mutation analysis. In the inactive, monomeric conformation the interaction between JH2 and the kinase domain (JH1) in part closes the JAK2 structure and hinders the transphosphorylation of the active kinase domains. The active conformation is a more open, dimeric structure formed partly via JH2-JH2 interactions between the opposing JAK2 proteins.

The model also gives theoretical background to our observation that ATP-binding to JH2 is crucial for the pathogenic JAK activation. It implies that the ATP-binding site is structurally important region within the pseudokinase that directly affects to the dynamic shifting between the active and inactive JH2 conformations.

The results summarized above allow us to better comprehend the characteristics that dictate the specificity among JAK-signaling pathways. Moreover, they bring insight into the mechanism of pathologic JAK activation and support the development of novel, more potent JAK inhibitors.

The doctoral dissertation of M.Sc. Juuli Raivola in the field of molecular immunology titled Molecular Regulation of Janus Kinases (JAKs) - Focus on the Pseudokinase Domain will be publicly examined at the Faculty of Medicine and Health Technology of Tampere University at 12 o'clock on Friday 22 January.  The opponent will be professor Stefan N. Constantinescu from Université catholique de Louvain, Belgium, while Professor Olli Silvennoinen will act as the custos.

The event can be followed via remote connection.

The dissertation is available online at
http://urn.fi/URN:ISBN:978-952-03-1828-4