scholarly journals Phospholipase C–mediated hydrolysis of PIP2 releases ERM proteins from lymphocyte membrane

2009 ◽  
Vol 184 (3) ◽  
pp. 451-462 ◽  
Author(s):  
Jian-Jiang Hao ◽  
Yin Liu ◽  
Michael Kruhlak ◽  
Karen E. Debell ◽  
Barbara L. Rellahan ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Phospholipase C ◽  
Membrane Association ◽  
Erm Proteins ◽  
Cytoskeletal Remodeling ◽  
In Vitro Binding ◽  
Vitro Binding ◽  
Hydrolysis Of

Mechanisms controlling the disassembly of ezrin/radixin/moesin (ERM) proteins, which link the cytoskeleton to the plasma membrane, are incompletely understood. In lymphocytes, chemokine (e.g., SDF-1) stimulation inactivates ERM proteins, causing their release from the plasma membrane and dephosphorylation. SDF-1–mediated inactivation of ERM proteins is blocked by phospholipase C (PLC) inhibitors. Conversely, reduction of phosphatidylinositol 4,5-bisphosphate (PIP2) levels by activation of PLC, expression of active PLC mutants, or acute targeting of phosphoinositide 5-phosphatase to the plasma membrane promotes release and dephosphorylation of moesin and ezrin. Although expression of phosphomimetic moesin (T558D) or ezrin (T567D) mutants enhances membrane association, activation of PLC still relocalizes them to the cytosol. Similarly, in vitro binding of ERM proteins to the cytoplasmic tail of CD44 is also dependent on PIP2. These results demonstrate a new role of PLCs in rapid cytoskeletal remodeling and an additional key role of PIP2 in ERM protein biology, namely hydrolysis-mediated ERM inactivation.

scholarly journals Morphogenic Effects of Ezrin Require a Phosphorylation-Induced Transition from Oligomers to Monomers at the Plasma Membrane

2000 ◽  
Vol 150 (1) ◽  
pp. 193-204 ◽  
Author(s):  
Alexis Gautreau ◽  
Daniel Louvard ◽  
Monique Arpin
Keyword(s):  
Plasma Membrane ◽  
Epithelial Cells ◽  
Wild Type ◽  
Erm Proteins ◽  
Threonine Residue ◽  
Phosphorylation Event ◽  
Membrane Bound

ERM (ezrin, radixin, moesin) proteins act as linkers between the plasma membrane and the actin cytoskeleton. An interaction between their NH2- and COOH-terminal domains occurs intramolecularly in closed monomers and intermolecularly in head-to-tail oligomers. In vitro, phosphorylation of a conserved threonine residue (T567 in ezrin) in the COOH-terminal domain of ERM proteins disrupts this interaction. Here, we have analyzed the role of this phosphorylation event in vivo, by deriving stable clones producing wild-type, T567A, and T567D ezrin from LLC-PK1 epithelial cells. We found that T567A ezrin was poorly associated with the cytoskeleton, but was able to form oligomers. In contrast, T567D ezrin was associated with the cytoskeleton, but its distribution was shifted from oligomers to monomers at the membrane. Moreover, production of T567D ezrin induced the formation of lamellipodia, membrane ruffles, and tufts of microvilli. Both T567A and T567D ezrin affected the development of multicellular epithelial structures. Collectively, these results suggest that phosphorylation of ERM proteins on this conserved threonine regulates the transition from membrane-bound oligomers to active monomers, which induce and are part of actin-rich membrane projections.

Localization of “in vitro” binding of the fungal toxin fusicoccin to plasma-membrane-rich fractions from corn coleoptiles

1977 ◽  
Vol 9 (3) ◽  
pp. 291-299 ◽  
Author(s):  
U. Dohrmann ◽  
R. Hertel ◽  
P. Pesci ◽  
S.M. Cocucci ◽  
E. Marrè ◽  
...  
Keyword(s):  
Plasma Membrane ◽  
Fungal Toxin ◽  
In Vitro Binding ◽  
Vitro Binding

scholarly journals Dynamics of Ankyrin-containing Complexes in Chicken Embryonic Erythroid Cells: Role of Phosphorylation

2001 ◽  
Vol 12 (12) ◽  
pp. 3864-3874 ◽  
Author(s):  
Sourav Ghosh ◽  
John V. Cox
Keyword(s):  
Complex Formation ◽  
Anion Exchanger ◽  
Erythroid Cells ◽  
Binding Studies ◽  
Phosphatase Inhibitors ◽  
In Vitro Binding ◽  
Vitro Binding

Chicken erythroid ankyrin undergoes a fairly rapid cycle of cytoskeletal association, dissociation, and turnover. In addition, the cytoskeletal association of ankyrin is regulated by phosphorylation. Treatment of erythroid cells with serine and threonine phosphatase inhibitors stimulated the hyperphosphorylation of the 225- and 205-kDa ankyrin isoforms, and dissociated the bulk of these isoforms from cytoskeletal spectrin. In vitro binding studies have shown that this dissociation of ankyrin from spectrin in vivo can be attributed to a reduced ability of hyperphosphorylated ankyrin to bind spectrin. Interestingly, a significant fraction of detergent insoluble ankyrin accumulates in a spectrin-independent pool. At least some of this spectrin-independent pool of ankyrin is complexed with the AE1 anion exchanger, and the solubility properties of this pool are also regulated by phosphorylation. Treatment of cells with serine and threonine phosphatase inhibitors had no effect on ankyrin/AE1 complex formation. However, these inhibitors were sufficient to shift ankyrin/AE1 complexes from the detergent insoluble to the soluble pool. These analyses, which are the first to document the in vivo consequences of ankyrin phosphorylation, indicate that erythroid ankyrin-containing complexes can undergo dynamic rearrangements in response to changes in phosphorylation.

scholarly journals The Development of Novel In Vitro Binding Assays to Further Elucidate the Role of tRNAs in Protein Synthesis

10.5772/21822 ◽  
2011 ◽  
Author(s):  
Anthony J. ◽  
Jr. ◽  
Suzanna Ellzey ◽  
Devin McDougald ◽  
Crystal Serrano
Keyword(s):  
Protein Synthesis ◽  
Binding Assays ◽  
In Vitro Binding ◽  
Vitro Binding
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