G Catenin and actin cytoskeleton dynamics

Actin mediated changes in cell shape, are essential for a wide range of cellular activities, from cell motility to dendritogenesis to axon guidance.

The actin binding properties of some Catenins together with their strategic proximity to the cellular membrane and its molecular machinery confer to these molecules important roles in the regulation of cytoskeletal dynamics in response to extracellular stimuli. All known phenotypes of GCatenin, from the effect on cell junctions in MCDK cells to the effect in dendritic organization in hippocampal neurons are very likely to be exerted via a G–Catenin mediated change in cytoskeletal processes.

GCatenin has been shown to physically interact with actin in cell lines and cultured hippocampal neurons where they co-localize in growth cones (Lu et al., 2002).

GCatenin also interacts with cortactin. Cortactin is a linker protein in the actin cytoskeleton: as it cross-links actin filaments in a tyrosine phosphorylation dependent manner (Weed and Parson 1993, Huang et al., 1997).GCatenin and cortactin form a complex in which a COOH region just downstream of the last Arm repeat of G-Catenin appears to be crucial (Martinez et al., 2003). In rat hippocampal neurons and in PC12 the complex is responsible for primary process extension and its absence (by deleting G- Catenin and Cortactin interaction domains) has negative effects on the ability of the cells to generate processes and branches. The G-Catenin and cortactin complex recruits the Arp 2/3 complex through which the actin dependent-outgrowth process is most likely driven. The Arp 2/3 complex comprises 7 polypeptides and regulates both the formation and structure of actin networks directly. By increasing the nucleation rate, the Arp2/3 complex generates the large number of new filaments needed for actin network formation and helps create the branched network by cross-linking the slow growing pointed end of one filament to the side of another (May, 2001; Weaver et al., 2001). The Arp 2/3 complex-mediated cross-links are relatively unstable and cortactin and G-Catenin may

stabilize the Arp 2/3 complex-mediated branches. This may serve to localize protrusions to sites of neuronal activity in light of the interaction of both G-Catenin and Cortactin with postsynaptic scaffolding proteins. The interaction between GCatenin, Cortactin and in turn Arp 2/3 is regulated by the state of phosphorylation of GCatenin and Cortactin. The application H2O2 and orthovanadate triggers GCatenin phosphorylation, via a non identified Src kinase family member and causes the dissociation of the complex and the inhibition of processes outgrowth (Martinez et al., 2003).

Process elongation and process branching are most likely regulated in a different fashion, the modulation of neurite complexity being a result of the balance between the two. As mentioned before, Rho A has been shown to be important in dendritic branching. Its inhibition, in particular, promotes dendritic branching whereas its over-expression or the expression of dominant active versions has strong inhibitory effects. The generation of new primary processes and their elongations are, on the other hand, mostly unaffected (Nakayama et al., 2000 and Neumann et al., 2002). GCatenin appears to be connected with both process elongation and process branching as its over-expression enhances (or mimics) the effects of Rho A mutants (or Rho A inhibitors) through a possible G-Catenin Rho A inhibitory activity. This still unidentified mechanism may be similar to that of its close relative p120ctn (Anastasiadis et al., 2000 and Noren et al., 2000). On the other hand,GCatenin mutants unable to form a complex with cortactin show a decrease in the number of primary dendrites and a defect in their elongation whereas the Rho A- dependent branching activity is partially retained (Martinez et al., 2003) (See figure 5.18)

Figure 5.18. GCatenin and primary process extension versus branching. Two different pathways regulate the effects of GCatenin on process elaboration. UnphosphorylatedGCatenin forms a complex with unphosphorylated Cortactin; the complex then recruits Arp2/3 to promote new generation of actin filaments. Not yet identified extracellular signals can lead to phosphorylation of theG-Catenin–Cortactin complex causing the disruption of their interaction and the complex with Arp2/3 arresting dendrite elongation. Rho inhibition can be amplified by phosphorylatedG-Catenin, which leads to branching (modified from Martinez et al., 2003).

G-Catenin has also been shown to bind the cytoplasmic non receptor tyrosine kinase c- Abl (the cellular homologue of Abelson murine leukemia virus) (Lu et al., 2002). The presence of nuclear and cytoplasmic pools of Abl and its actin-binding capability indicates a role of the molecule in the regulation of cell cycle, cytoskeletal organization (Van Etten, 1999) and in neuronal morphogenesis (Koleske et al., 1998 and Zuckerberg et al., 2000). Abl interacts with several different protein families, including catenin/cadherin cell adhesion complexes, Trio family GFP exchange factors (GEFs) and

Ena/VASP (vasodilator-stimulated phosphor-protein) family actin regulatory proteins (Lanier and Gertler, 2000).

GCatenin has strong consensus sites for Abl binding and Abl-induced tyrosine phosphorylation in the N terminal part of the molecule. The two proteins form a stable complex in neurons and they co localize with F-actin filaments in the growth cone. Furthermore MDCK cells irradiated with UV light (which was shown to activate c-Abl kinase activity) shows a strong GCatenin phosphorylation. The use of specific c-Abl kinase inhibitors enhances the effects of G-Catenin in PC12 differentiation upon treatment with NGF (Lu et al., 2002). The alteration of MDCK cell shape in response to HGF (Lu et al., 1999) is also more pronounced in the presence of c-Abl kinase inhibitors. Both events are mediated by a reorganization of the actin cytoskeleton, consistently with the hypothesis that the unphosphorylatedG-Catenin is the one responsible for actin polymerization promoting/regulating activity.