Beta1PIX, the PAK-interacting exchange factor, requires localization via a coiled-coil region to promote microvillus-like structures and membrane ruffles
Glaxo-IMCB Group, Institute of Molecular and Cell Biology, 30 Medical Drive, Singapore 117609.Journal of Cell Science (Impact Factor: 5.43). 01/2002; 114(Pt 23):4239-51.
PIX is a Rho-family guanine nucleotide exchange factor that binds PAK. We previously described two isoforms of PIX that differ in their N termini. Here, we report the identification of a new splice variant of betaPIX, designated beta2PIX, that is the dominant species in brain and that lacks the region of approximately 120 residues with predicted coiled-coil structure at the C terminus of beta1PIX. Instead, beta2PIX contains a serine-rich C terminus. To determine whether these splice variants differ in their cellular function, we studied the effect of expressing these proteins in HeLa cells. We found that the coiled-coil region plays a key role in the localization of beta1PIX to the cell periphery and is also responsible for PIX dimerization. Overexpression of beta1, but not beta2PIX, drives formation of membrane ruffles and microvillus-like structures (via activation of Rac1 and Cdc42, respectively), indicating that its function requires localized activation of these GTPases. Thus, beta1PIX, like other RhoGEFs, exerts specific morphological functions that are dependent on its intracellular location and are mediated by its C-terminal dimerization domain.
[Show abstract] [Hide abstract]
- "The GIT–PIX complex is multimeric. GIT proteins form homodimers and heterodimers through their coiled coil domains (Paris et al., 2003; Premont et al., 2004), likewise similar domains in the C-terminus of β-PIX enable its homodimerisation (Kim et al., 2001; Koh et al., 2001). Recent structural studies have suggested that β-PIX forms trimers rather than dimers, resulting in a heteropentameric complex in which each SHD of GIT1 binds the GIT-binding domains of two of the three β-PIX proteins (Schlenker and Rittinger, 2009). "
ABSTRACT: RhoJ is a RhoGTPase expressed in endothelial cells and tumour cells which regulates cell motility, invasion, endothelial tube formation and focal adhesion numbers. This study aimed to further delineate the molecular function of RhoJ. Using timelapse microscopy RhoJ was found to regulate focal adhesion disassembly; siRNA-mediated knockdown of RhoJ increased focal adhesion disassembly time, while expression of an active mutant (daRhoJ) decreased it. Further, daRhoJ co-precipitated with the GIT-PIX complex, a regulator of focal adhesion disassembly. An interaction between daRhoJ and GIT1 was confirmed using yeast-2-hybrid, which depended on the Spa homology domain of GIT1. GIT1, GIT2, β-PIX and RhoJ all co-localised in focal adhesions and depended on each other for their recruitment to focal adhesions. Functionally, the GIT-PIX complex regulated endothelial tube formation, with knockdown of GIT1/2 or β-PIX phenocopying RhoJ knockdown. RhoJ knockout mice showed reduced tumour growth and diminished tumour vessel density, identifying a role for RhoJ in mediating tumour angiogenesis. These studies give novel insight into the molecular function of RhoJ in regulating cell motility and tumour vessel formation.Journal of Cell Science 06/2014; 127(14). DOI:10.1242/jcs.140434 · 5.43 Impact Factor
[Show abstract] [Hide abstract]
- "For instance, β-catenin binds to scribble, a tumor suppressor factor implicated in epithelial cell polarity and in synapse formation , , . Scribble binds to β-Pix, a guanine nucleotide exchange factor (GEF) for Rho GTPases , and with its partner GIT1, a GTPase-activating protein (GAP) for the ARF family of small GTPases –. Scribble controls the internalization of hormone receptors by regulating ARF6 via the β-Pix-GIT1 complex , and activation of ARF6 and Rac1 in CHO cells expressing muscular nAChRs causes nAChRs endocytosis , . "
ABSTRACT: Neuronal nicotinic acetylcholine receptors (nAChRs) are widely expressed throughout the central and peripheral nervous system and are localized at synaptic and extrasynaptic sites of the cell membrane. However, the mechanisms regulating the localization of nicotinic receptors in distinct domains of the cell membrane are not well understood. N-cadherin is a cell adhesion molecule that mediates homotypic binding between apposed cell membranes and regulates the actin cytoskeleton through protein interactions with the cytoplasmic domain. At synaptic contacts, N-cadherin is commonly localized adjacent to the active zone and the postsynaptic density, suggesting that N-cadherin contributes to the assembly of the synaptic complex. To examine whether N-cadherin homotypic binding regulates the cell surface localization of nicotinic receptors, this study used heterologous expression of N-cadherin and α3β4 nAChR subunits C-terminally fused to a myc-tag epitope in Chinese hamster ovary cells. Expression levels of α3β4 nAChRs at cell-cell contacts and at contact-free cell membrane were analyzed by confocal microscopy. α3β4 nAChRs were found distributed over the entire surface of contacting cells lacking N-cadherin. In contrast, N-cadherin-mediated cell-cell contacts were devoid of α3β4 nAChRs. Cell-cell contacts mediated by N-cadherin-deleted proteins lacking the β-catenin binding region or the entire cytoplasmic domain showed control levels of α3β4 nAChRs expression. Inhibition of actin polymerization with latrunculin A and cytochalasin D did not affect α3β4 nAChRs localization within N-cadherin-mediated cell-cell contacts. However, treatment with the Rho associated kinase inhibitor Y27632 resulted in a significant increase in α3β4 nAChR levels within N-cadherin-mediated cell-cell contacts. Analysis of α3β4 nAChRs localization in polarized Caco-2 cells showed specific expression on the apical cell membrane and colocalization with apical F-actin and the actin nucleator Arp3. These results indicate that actomyosin contractility downstream of N-cadherin homotypic binding regulates the cell surface localization of α3β4 nAChRs presumably through interactions with a particular pool of F-actin.PLoS ONE 04/2013; 8(4):e62435. DOI:10.1371/journal.pone.0062435 · 3.23 Impact Factor
[Show abstract] [Hide abstract]
- "Besides the cc-domain, the pleckstrin homology (PH) domain of PIX proteins has also been reported to play a role in intracellular targeting. As the PH domain of aPIX and bPIX share only 72% amino acid homology (Koh et al., 2001), they might be directed to different target sites within the same cell. Importantly, aPIX and bPIX may also form heterodimers via their cc-domains (Rosenberger et al., 2003). "
ABSTRACT: In the cerebral cortex of reeler mutant mice lacking reelin expression, neurons are malpositioned and display misoriented apical dendrites. Neuronal migration defects in reeler have been studied in great detail, but how misorientation of apical dendrites is related to reelin deficiency is poorly understood. In wild-type mice, the Golgi apparatus transiently translocates into the developing apical dendrite of radially migrating neurons. This dendritic Golgi translocation has recently been shown to be promoted by reelin. However, the underlying signalling mechanisms are largely unknown. Here, we show that the Cdc42/Rac1 guanine nucleotide exchange factor αPIX/Arhgef6 promoted translocation of Golgi cisternae into developing dendrites of hippocampal neurons. Reelin treatment further increased the αPIX-dependent effect. In turn, overexpression of exchange activity-deficient αPIX or dominant-negative (dn) Cdc42 or dn-Rac1 impaired dendritic Golgi positioning, an effect that was not compensated by reelin treatment. Together, these data suggest that αPIX may promote dendritic Golgi translocation, as a downstream component of a reelin-modulated signalling pathway. Finally, we found that reelin promoted the translocation of the Golgi apparatus into the dendrite that was most proximal to the reelin source. The distribution of reelin may thus contribute to the selection of the process that becomes the apical dendrite.European Journal of Neuroscience 02/2013; 37(9). DOI:10.1111/ejn.12153 · 3.18 Impact Factor
Data provided are for informational purposes only. Although carefully collected, accuracy cannot be guaranteed. The impact factor represents a rough estimation of the journal's impact factor and does not reflect the actual current impact factor. Publisher conditions are provided by RoMEO. Differing provisions from the publisher's actual policy or licence agreement may be applicable.