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Regulation of Protein Tyrosine Kinase 6 Stability and Activation
thesisposted on 2022-05-01, 00:00 authored by Milica B. Gilic
Protein Tyrosine Kinase 6 (PTK6) is an intracellular tyrosine kinase whose functions depend on its localization. When localized at the plasma membrane in prostate cancers, PTK6 promotes oncogenic signaling and is associated with worse patient outcomes. PTK6 acts growth inhibitory when it is localized in the nucleus. PTK6 lacks membrane targeting domains and nuclear localization sequences, and mechanisms of regulation of intracellular localization are not well understood. The SH2 domain of PTK6 was identified as crucial for the interaction with plasma membrane lipids which was shown to be mediated by two arginine residues, R131 and R136. Mutation of these residues did not lead to reduction in lipid binding. We show that PTK6 preferentially binds monophosphorylated phosphoinositides. Mutation of R131 and R136 led to increase in PTK6 stability. In contrast, mutation of amino acid residues R85 and H126 which are involved in binding of phosphotyrosines, led to accumulation of PTK6 in cellular granules and reduced stability. We demonstrate that wild type PTK6 and phosphotyrosine binding mutant interact with Cul1 and SKP1, which are components of Cullin Ring Ligase 1 E3 ubiquitin ligase complex. The more stable previously described as lipid binding mutant, does not interact with this complex. Mutation of PTK6 amino residue R131 has been reported in some cancers, which may contribute to increased PTK6 stability, activity, and cancer signaling. We show that activity of PTK6 is inhibited by Vemurafenib, in vitro and in vivo. Vemurafenib reduces proliferation and increases cell death in xenograft tumors, which suggests that PTK6 could be targeted in cancers. I found that calcium-binding protein, calmodulin can bind PTK6 and increase its autophosphorylation, which could be one of the pathways PTK6 is activated in differentiation of keratinocytes upon increase in calcium ion levels.
AdvisorTyner, Angela L.
ChairTyner, Angela L.
DepartmentBiochemistry and Molecular Genetics
Degree GrantorUniversity of Illinois at Chicago
Degree namePhD, Doctor of Philosophy
Committee MemberGaponenko, Vadim Hay, Nissim Raychaudhuri, Pradip Simonovic, Miljan Naba, Alexandra
Submitted dateMay 2022