We first described several years ago the finding that chemokine (SDF-1) stimulation of lymphocytes induces very rapid (onset <=5 sec) ERM dephosphorylation and inactivation. Given the evidence that phosphorylation contributes to ERM activation, it was plausible that dephosphorylation mediated the inactivation. However, membrane phospho-lipid, especially PIP2, is an additional key mediator of ERM activation. We hypothesized that acute reduction of PIP2 could contribute to ERM inactivation. Specifically, we considered the possibility that chemokine: 1) activates phospholipase C (PLC);which 2) mediates reduction of phosphatidylinositol 4,5-bisphosphate (PIP2);which 3) releases ERM from its association with the plasma membrane;and 4) triggers ERM dephosphorylation. This proposed model is supported by multiple findings we have made, including the following. PLC inhibitor blocks these events. Translocation of GFP-tagged moesin into cytosol and ERM dephosphorylation can be induced either by: transfection with active PLC construct;or by acute targeting of phosphoinositide 5-phosphatase to the plasma membrane. PIP2 dependence cannot be replaced by phosphorylation (modeled by T558D mutation) in either of two assays: association with plasma membrane in cells or in vitro association with cytoplasmic tails of proteins like CD44. Moreover, induction of PIP2 hydrolysis is sufficient to induce ERM dephosphorylation, indicating that PLC activation can be sufficient to induce ERM dephosphorylation and that PLC activation may be the initiating event in lymphocytes rather than independent activation of an ERM phosphatase. Key findings made with moesin constructs have been confirmed with ezrin constructs, indicating that these are general principles pertinent to both ezrin and moesin. These results demonstrate a powerful new role of PLCs in rapid cytoskeletal remodeling and an additional key role of PIP2 in ERM biology, namely hydrolysis-mediated ERM inactivation. These studies are described in a paper published this year in Journal of Cell Biology. The complex N-terminal domain of ERM is called the FERM domain which binds to the C-terminus and thereby participates in autoinhibition. In our view, the release of ERM autoinhibition by acidic phospholipid binding is complex and incompletely understood. Therefore, we are conducting additional mutational analysis of FERM domain residues and analyzing the mutant proteins for protein-protein interaction and membrane localization to extend our functional understanding of these key processes.
| Hao, Jian-Jiang; Liu, Yin; Kruhlak, Michael et al. (2009) Phospholipase C-mediated hydrolysis of PIP2 releases ERM proteins from lymphocyte membrane. J Cell Biol 184:451-62 |
