Calmodulin Binds to and Inhibits H‐Ras Activation of PI3K: A Single Molecule Study

Abstract

The lipid‐anchored GTPase Ras is well known for its role in oncogenesis, with around 25% of human tumors showing mutations in a Ras family member. One of the best characterized Ras effectors is the highly oncogenic lipid kinase phosphatidylinositol 3‐kinase (PI3K). PI3K phosphorylates the constitutive plasma membrane lipid PIP2, yielding the vital second messenger lipid PIP3. This PIP3 signal regulates many cell processes including migration and growth. Calmodulin also activates PI3K by tightly binding MARCKS, a PIP2 sequestering protein, freeing the lipid. The enhanced free PIP2 population serves as additional substrate for PI3K, increasing its activity. The findings presented here reveal that surprisingly, Ras activation of PI3K is blocked by the presence of calmodulin, with important implications for PIP3 signaling. We are observing these mechanisms using single molecule TIRF microscopy to probe the interactions and activities of single proteins on a supported target membrane surface. Preliminary evidence monitoring PI3K binding and lipid kinase activity at the single molecule level reveals that biologically relevant concentrations of calmodulin strongly inhibit Ras activation of PI3K. These results imply a new and additional level of control for an important and oncogenic signaling pathway.Support or Funding InformationFunding for this work was provided by the National Institutes of Health ( R01 GM063235 to J.J.F.), the Medical Research Council ( MC U105184308 to R.L.W.), the AstraZeneca/LMB Blue Skies Initiative ( MC A024‐5PF9G to R.L.W.), and St. Catharine's College (G.R.M.).This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.