PI5P4Kβ is a sensor of GTP concentration that activates lipid second messenger signaling.
Major finding: PI5P4Kβ is a sensor of GTP concentration that activates lipid second messenger signaling.
Mechanism: PI5P4Kβ preferentially uses GTP over ATP to phosphorylate PI(5)P to produce PI(4,5)P2.
Impact: GTP sensing by PI5P4Kβ spurs tumorigenesis, and may be a potential therapeutic target in cancer.
ATP and GTP levels vary in different tissue types and under different conditions, and concentration changes can be detected by sensor molecules. These sensors can bind to the metabolite, have a KM that allows its activity to be regulated by changes in metabolite concentration, and can modulate cellular functions. Although ATP sensors have been identified, GTP sensors remain unknown. To identify candidate GTP sensors, Sumita, Lo, Takeuchi, and colleagues performed a screen for GTP-binding proteins, and identified the phosphatidylinositol 5-phosphate 4-kinases (PI5P4K). PI5P4Kβ was determined to be a good candidate GTP sensor, as it bound directly to GTP more strongly than ATP. Excess ATP could compete with GTP for binding, indicating that GTP bound to the same site as ATP, but PI5P4Kβ was able to hydrolyze GTP five times faster than ATP. In vitro kinase assays demonstrated that, under physiologically relevant conditions, PI5P4Kβ utilized GTP to phosphorylate its substrate, the lipid second messenger PI(5)P, with approximately half of the resulting PI(4,5)P2 produced by using GTP. X-ray crystallography structures of PI5P4Kβ in complex with non-hydrolyzable GTP and ATP analogues indicated that GTP binds to the PI5P4Kβ hydrophobic groove, and were used to engineer a PI5P4Kβ mutant (PI5P4Kβ F205L) that disrupted GTP binding, but maintained ATP binding. The PI5P4KβF205L mutant lost its GTP-sensing ability, and cells expressing PI5P4KβF205L had higher levels of PI(5)P. PI5P4KβF205L expression resulted in formation of fewer colonies in anchorage-independent growth assays, compared with wild-type cells, and impaired tumor formation in in vivo allograft assays. These findings indicate that PI5P4Kβ is an intracellular GTP sensor that acts via phosphorylation of PI(5)P to regulate cell metabolism, and suggest that the GTP-sensing function of PI5P4Kβ promotes tumor growth in vivo and may be exploitable in the development of cancer therapeutics.
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