Plasma Membrane PI(4,5)P2 Threshold Regulates Chemotactic signaling pathways and cell morphology

Abstract

During chemotaxis, cytoskeletal and signaling biosensors show a polarized distribution on the plasma membrane (PM) by which support a role for a PM phosphatidylinositol 4,5-bisphosphate [PI(4,5)P2] gradient from the rear of the cell to the leading edge. We have found that the activity of the GTPase Ras is reciprocally regulated with the localization of the tumor suppressor PTEN. These studies suggest that PI(4,5)P2 levels set up a threshold for the activity of signaling molecules at the rear and the leading edge of a cell. To further demonstrate the importance of PM PI(4,5)P2 in maintaining cell morphology and breaking symmetry to establish a polarized morphology, we used a Dictyostelium mutant strain lacking the type I PIP5K, which is the main enzyme synthesizing PI(4,5)P2. These cells, designated pikI-, have highly reduced PI(4,5)P2 levels and higher Ras activity compared to wild-type cells. We found that pikI-resemble similar cell responses observed when we use an inducible system to deplete PM PI(4,5)P2 levels. Leading edge biosensors diffuse to the cytosol when the pikI- round-up and translocate back to the PM when the cells spread. These observations suggest that PI(4,5)P2 levels go down as cells spread, and elevate as cells round-up and contract. F-actin polymerization also occurs as the cell spread. Remarkably, Ras and PI3K biosensors are activated on the PM when actin polymerization is inhibited with Latrunculin A (LatA). The lower levels of PM PI(4,5)P2 in pikI- cells is likely below the minimal level of PI(4,5)P2 needed to inhibit Ras activity, and thus contributes to the turning on of signaling networks. We also tested the localization of the mDia-like formin A (for A) which functions to generate subsets of actin filaments at the cell's back likely via it's binding to the PM PI(4,5)P2. For A biosensor dynamics in pikI- showed a reciprocal localization with PH-RFP, a marker for PI(3,4,5)P3. Altogether, plasma membrane levels of PI(4,5)P2 are suggested to be critical in the cell morphology observed. Lower levels of PI(4,5)P2 contribute to activating excitatory networks controlling cell protrusions. PM PI(4,5)P2 levels above a threshold block front responses and support regulators of pathways leading to actomyosin contraction. Support or Funding Information NIH and W. W. Smith Charitable Trust This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.