Expression level, subcellular distribution and Rho-GDI binding affinity of merlin in comparison with ezrin/radixin/moesin proteins

Abstract

Merlin, a neuro®bromatosis type-2 tumor suppressor, shows signi®cant sequence similarity to ERM (Ezrin/ Radixin/Moesin) proteins, general actin ®lament/plasma membrane cross-linkers, which are regulated in a Rhodependent manner. To understand its physiological functions, we compared merlin with ERM proteins in vivo and in vitro. Quantitative immunoblotting revealed that the molar ratio of merlin/ERM in cultured epithelial or non-epithelial cells was *0.14 or *0.05, respectively. After centrifugation of cell homogenate, merlin was mostly recovered in the insoluble fraction, whereas almost half of ERM proteins were found in the soluble fraction. Merlin and ERM proteins were concentrated at microvilli when introduced into ®broblasts. In contrast, in epithelial cells, introduced merlin was co-distributed with E-cadherin in lateral membranes, whereas ERM proteins were concentrated in apical microvilli. Finally, we examined the binding anity of merlin to Rho GDP dissociation inhibitor (Rho-GDI), to which N-terminal halves of ERM proteins but not the full-length molecules speci®cally bind. In vitro binding assays revealed that the N-terminal halves of merlin isoform-I and -II as well as full-length merlin isoform-II bound to Rho-GDI with similar binding anity to ERM proteins. Immunoprecipitation con®rmed these ®ndings in vivo. These ®ndings do not favor the notion that merlin functions simply in a redundant or competitive manner to ERM proteins.

Cortical Actin Organization: Lessons from ERM (Ezrin/Radixin/Moesin) Proteins

“…The subcellular localization of ERM proteins was similar to that of merlin in fibroblasts (microvilli and ruffling membranes) but different in epithelial cells; merlin, but not ERM proteins, was concentrated at lateral membranes together with Ecadherin (64). The N-terminal half of merlin bound to the cytoplasmic domains of CD44 (65) and NHE-RF (66) as well as Rho-GDI in vitro (64).…”

Magicin, a novel cytoskeletal protein associates with the NF2 tumor suppressor merlin and Grb2

“…Merlin interacts with F-actin through actin binding sites within the FERM domain (Xu and Gutmann, 1998;Brault et al, 2001;James et al, 2001). In addition to actin, several other merlin interacting proteins have been identified, which include b-fodrin/bII-spectrin (Scoles et al, 1998;Neill and Crompton, 2001), SCHIP-1 (Goutebroze et al, 2000), NHE-RF (Murthy et al, 1998), b1-integrin (Obremski et al, 1998), CD44 (Sainio et al, 1997;Morrison et al, 2001), HRS (Scoles et al, 2000), RhoGDI (Maeda et al, 1999), syntenin (Jannatipour et al, 2001), paxillin (Fernandez-Valle et al, 2002) and RIb subunit of the cAMP-protein kinase A (Gronholm et al, 2003). Among these proteins, NHE-RF, CD44 and RhoGDI have been independently identified as ERM binding partners (Tsukita et al, 1994;Reczek et al, 1997;Takahashi et al, 1997), while HRS and syntenin appear to be specific for merlin.…”

Temporal Control of Rac in Schwann Cell–Axon Interaction Is Disrupted inNF2-Mutant Schwannoma Cells

“…Schwannoma cells have increased active Rac and its downstream effector p21-activated kinase (PAK) (Kaempchen et al, 2003). NF2 is involved in suppression of Rac-PAK signaling by blocking the p21 binding domain (PBD) of PAK (Kissil et al, 2003) or by binding to Rho-GDI, a GDP dissociation inhibitor, that may stabilize the inactive form of Rac (Maeda et al, 1999). Loss of NF2 alters Rac-PAK-mediated proliferation (Xiao et al, 2005), migration (Shaw et al, 2001), and contact inhibition (Okada et al, 2005).…”