Phosphatidylinositol (4,5) Bisphosphate Controls T Cell Activation by Regulating T Cell Rigidity and Organization

Abstract

Here we investigate the role of Phosphatidylinositol (4,5) bisphosphate (PIP2) in the physiological activation of primary murine T cells by antigen presenting cells (APC) by addressing two principal challenges in PIP2 biology. First, PIP2 is a regulator of cytoskeletal dynamics and a substrate for second messenger generation. The relative importance of these two processes needs to be determined. Second, PIP2 is turned over by multiple biosynthetic and metabolizing enzymes. The joint effect of these enzymes on PIP2 distributions needs to be determined with resolution in time and space. We found that T cells express four isoforms of the principal PIP2-generating enzyme phosphatidylinositol 4-phosphate 5-kinase (PIP5K) with distinct spatial and temporal characteristics. In the context of a larger systems analysis of T cell signaling, these data identify the T cell/APC interface and the T cell distal pole as sites of differential PIP2 turnover. Overexpression of different PIP5K isoforms, as corroborated by knock down and PIP2 blockade, yielded an increase in PIP2 levels combined with isoform-specific changes in the spatiotemporal distributions of accessible PIP2. It rigidified the T cell, likely by impairing the inactivation of Ezrin Moesin Radixin, delayed and diminished the clustering of the T cell receptor at the cellular interface, reduced the efficiency of T cell proximal signaling and IL-2 secretion. These effects were consistently more severe for distal PIP5K isoforms. Thus spatially constrained cytoskeletal roles of PIP2 in the control of T cell rigidity and spatiotemporal organization dominate the effects of PIP2 on T cell activation.

Platelets lacking PIP5KIγ have normal integrin activation but impaired cytoskeletal-membrane integrity and adhesion

“…These data are consistent with work by Raucher et al 45,46 that demonstrated that marked disruption of PIP 2 by overexpression of phospholipidsequestering polypeptides or inositol phosphatases or by pharmacologic activation of phospholipase C reduces the adhesion energy between the cell membrane and the cytoskeleton. Our data also support the findings by Sun et al, 47 which showed that overexpression of PIP5KIg increased the rigidity of the membrane in T cells. The structural mediators of this PIP 2 -driven signal are less clear, but candidate PIP 2 -binding proteins are vinculin, talin, and filamin.…”

Vinculin Arrests Motile B Cells by Stabilizing Integrin Clustering at the Immune Synapse

“…They remain detectable at the pSMAC of the mature synapse, possibly assisting the active vinculin conformation. PIPKIg is also found at the T cell synapse; the spatiotemporal regulation of PIP 2 synthesis appears to control T cell rigidity and signaling organization (42).…”

Phosphatidylinositol 4-Phosphate 5-Kinase β Controls Recruitment of Lipid Rafts into the Immunological Synapse

“…In T lymphocytes, PIP2 concentrates at the IS at a very early stage upon Ag recognition (43), and data obtained by overexpressing PIP5K isoforms in transgenic mice showed the enrichment of distinct isoforms at the IS (20).…”

“…Primary CD4 + T cells express all three PIP5K isoforms (a, b, and g), with a specific subcellular localization (19). Human PIP5Ka accumulates in a sustained manner in the T:APC contact zone (20), where it regulates CD28-mediated actin polymerization events necessary for CD28 costimulatory signaling (21,22). PIP5Kg90 is predominantly found at the distal pole and in the uropod.…”

“…PIP5Kg90 is predominantly found at the distal pole and in the uropod. PIP5Kb and PIP5Kg87 are rapidly but transiently recruited to the site of T:APC contacts during IS formation (20). PIP5Kg87 has been implicated in the formation of a stable T:APC interaction by regulating the transition of LFA-1 from a low intermediate to a high-activation state (23).…”