Rho-dependent Regulation of Cell Spreading by the Tetraspan Membrane Protein Gas3/PMP22

Abstract

Gas3/PMP22 plays a crucial role in regulating myelin formation and maintenance, and different genetic alterations in gas3/PMP22 are responsible for a set of human peripheral neuropathies. We have previously demonstrated that Gas3/PMP22 could regulate susceptibility to apoptosis in NIH3T3 cells but not in REF 52 cells. In this report we demonstrate that when the apoptotic response triggered by gas3/PMP22 was counteracted by Bcl-2 coexpression, morphological changes were observed. Time-lapse analysis confirmed that Gas3/PMP22 can modulate cell spreading, and this effect was strengthened after inhibition of phosphoinositide 3-kinase. Using the active form of the small GTPase RhoA, we have been able to dissect the different Gas3/PMP22 biological activities. RhoA counteracted the Gas3/PMP22-dependent morphological response but was unable to neutralize the apoptotic response. Treatment of NIH3T3 cells with cytotoxic necrotizing factor 1, which activates endogenous Rho, also counteracted Gas3/PMP22-mediated cell shape and spreading changes. Treatment of REF 52 cells, which are unresponsive to Gas3/PMP22 overexpression, with the C3 exoenzyme, inhibiting Rho activity, renders REF 52 cells responsive to Gas3/PMP22 overexpression for cell shape and spreading changes. Finally, assembly of stress fibers and focal adhesions complexes, in response to lysophosphatidic acid-induced endogenous Rho activation, was impaired in Gas3/PMP22-overexpressing cells. We hypothesize that cell shape and spreading regulated by Gas3/PMP22 through the Rho GTPase might have an important role during Schwann cells differentiation and myelinization.

Peripheral Myelin Protein 22 Is in Complex with α6β4 Integrin, and Its Absence Alters the Schwann Cell Basal Lamina

“…4 -7), together with our findings in an epithelial wound healing model (Roux et al, 2005), suggest a direct role for PMP22 in mediating the interactions between cells and the ECM. Cell-matrix interactions can influence cell shape, which is profoundly modulated by PMP22 (Brancolini et al, 1999;Roux et al, 2005). In fibroblasts and SCs, the overexpression of PMP22 induces membrane blebbing and regulates cell spreading (Brancolini et al, 2000), whereas in MDCK (Madin-Darby canine kidney) epithelial monolayers it leads to a flattened morphology (Roux et al, 2005).…”

“…The glycosylation consensus sequence at asparagine 41, which bears this epitope, is conserved among members of the PMP22 gene family and across species as distant as zebrafish (Taylor et al, 1995;Wulf et al, 1999). This carbohydrate modification of PMP22 is involved in the stabilization of PMP22 homodimers (Tobler et al, 1999) and in the cell spreading effects (Brancolini et al, 1999(Brancolini et al, , 2000, but not in the trafficking of the protein to the SC membrane (Ryan et al, 2000), or in the interaction with protein zero (Hasse et al, 2004). The L2/HNK-1 epitope can also modulate the heterophilic interaction of some nervous system molecules, including ␣1␤1 integrin and laminin (Pesheva et al, 1987;Lallier and BronnerFraser, 1992).…”

“…Overexpression of PMP22 in epithelial cells alters their migratory behavior and changes the functional properties of cell-cell contacts (Roux et al, 2005). In addition, in NIH3T3 cells (Brancolini et al, 1999) and rat SCs (Nobbio et al, 2004), the overexpression of PMP22 mediates membrane expansion. These findings, together with the pheno-types of PMP22 mutant mice, indicate that PMP22 is capable of modulating intercellular and cell-matrix communications in a variety of cell types.…”

Modulation of Epithelial Morphology, Monolayer Permeability, and Cell Migration by Growth Arrest Specific 3/Peripheral Myelin Protein 22

“…Conversely, reduced PMP22 mRNA levels are associated with enhanced DNA synthesis and entry into the S ϩ G2/M phases (Zoidl et al, 1995). In NIH 3T3 fibroblasts, PMP22 overexpression regulates cell spreading, an effect that is dependent on the Rho-GTPase pathway (Brancolini et al, 1999). Recent studies have detected exogenous PMP22 in ADP-ribosylation factor 6 (Arf-6)-positive plasma membrane-endosomal recycling vacuoles before apoptosis or changes in cell shape (Chies et al, 2003).…”

“…Transmembrane proteins of the apical junctional complex such as the claudins, occludin, and the junctional adhesion molecules (González-Mariscal et al, 2003) all participate in the regulation of junctional permeability. In addition, based on the findings of Brancolini and colleagues (Brancolini et al, 1999(Brancolini et al, , 2000Chies et al, 2003), PMP22 might be involved in the regulation of epithelial proliferation and/or cell migration, dynamic processes that involve changes in cell adhesion and morphology. In support of this possibility, PMP22 contains the carbohydrate L2/HNK1 adhesion/recognition epitope in the first extracellular loop (Snipes et al, 1993;Schachner et al, 1995).…”