Table of Contents

Volume 2, Issue 6

  November/December 2011

  Pages 293 - 333

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EDITOR'S CORNER

Open Access Article

In this issue of Small GTPases

Elaine L. Ellerton
Pages 293 - 294
http://dx.doi.org/10.4161/sgtp.19248
Full Text | PDF




RESEARCH PAPER

Open Access Article

Optimization and stabilization of Rho small GTPase proteins for solution NMR studies: The case of Rnd1

Shufen Cao and Matthias Buck
Pages 295 - 304
http://dx.doi.org/10.4161/sgtp.19257
Abstract | Full Text | PDF

Rho GTPases of the Ras superfamily have important roles in regulating the organization of the actin filament system, morphogenesis and migration of cells. Structural details for these proteins are still emerging, and information on their dynamics in solution is much needed to understand the mechanisms underlying their signaling functions. This report reviews conditions for solution NMR studies of Rho GTPases and describes our optimization and stabilization of Rnd1 for such experiments. Rnd1 belongs to the Rnd protein subfamily branch of Rho small GTPases and functions in neurite outgrowth, dendrite development and in axon guidance. However, as we report here, solution NMR studies of this protein are challenging. Multiple methods have been employed to enhance the stability of Rnd1, including by cleavage of an N-terminal His expression tag and by addition of non-hydrolysable GMPPNP (β: γ-imidoguanosine 5'-triphosphate) nucleotide. Further stabilization of Rnd1 against aggregation was achieved through a structure informed point mutation while maintaining its conformation and binding affinity for a partner protein. The NMR spectrum of the optimized protein reveals significant improvement in NMR signal dispersion and intensity. This work paves the way for structural and protein-protein/protein-ligand interaction studies of Rnd1 by solution NMR and also provides a guide for optimization and stabilization of other Rho GTPases.




EXTRA VIEWS

Open Access Article

Rac GTPase signaling in mechanotransduction during embryonic morphogenesis

Michel Labouesse
Pages 305 - 309
http://dx.doi.org/10.4161/sgtp.18035
Abstract | Full Text | PDF

How cells sense and respond to mechanical forces is attracting considerable attention. We recently demonstrated that mechanical tension originating from one tissue strongly influences the differentiation and morphogenesis of another tissue during C. elegans embryogenesis (Nature 471:99–103). Specifically, we found that the repeated contractions of muscle cells stimulate a signaling cascade involving the Rac GTPase within the epidermis. This pathway ultimately leads to strengthen hemidesmosome-like junctions and promote embryonic morphogenesis. Our work provides further evidence that mechanical inputs impact on development, much like inputs involving growth factors and morphogens. After briefly outlining the pioneering work that inspired us, I will present the mechanotransduction process underlying the response to tension and the key experiments supporting our conclusions.

Open Access Article

Atypical RhoV and RhoU GTPases control development of the neural crest

Sandrine Faure and Philippe Fort
Pages 310 - 313
http://dx.doi.org/10.4161/sgtp.18086
Abstract | Full Text | PDF

This review addresses the developmental roles of two GTPases of the Rho family, RhoV/Chp and RhoU/Wrch. These two GTPases form a distinct subfamily related to Rac and Cdc42 proteins and were detected in a screen for Rho members that are particularly expressed in the neural crest, an embryonic tissue peculiar to vertebrates. The neural crest represents a physiological model of normal epithelial to mesenchymal transition (EMT), in which epithelial cells at the border of neural and non-neural ectoderm differentiate, lose their intercellular connections and migrate throughout the embryo. We showed that RhoV, transiently induced by the canonical Wnt pathway, is required for the full differentiation of neural crest cells, while RhoU, induced later by the non-canonical Wnt pathway, is necessary for the migration process. These two GTPases, which are highly conserved across vertebrates, are thus tightly functionally linked to Wnt signaling, whose implication in embryonic development and cancer progression is well established. In the light of the recent literature, we discuss how RhoV and RhoU may achieve their physiological functions.

Open Access Article

Cellular mechanism of bile acid-accelerated hepatocyte polarity

Dong Fu, Jennifer Lippincott-Schwartz and Irwin M. Arias
Pages 314 - 317
http://dx.doi.org/10.4161/sgtp.18087
Abstract | Full Text | PDF

We recently discovered that the major mammalian bile acid, taurocholate, accelerated polarity in primary rat hepatocytes. Taurocholate increased cellular cAMP and signals through an Epac-Rap1-MEK-LKB1-AMPK pathway for its polarity effect. This review discusses possible mechanisms for how taurocholate affects different cell polarity factors, particularly AMPK, and thereby regulates events that generate polarity. These include tight junction formation, apical trafficking, recycling endosome dynamics, and cytoskeleton rearrangement. We also discuss whether the effects of taurocholate are mediated by other LKB1 downstream kinases, such as Par1 and NUAK1.

Open Access Article

Exploiting the origins of Ras mediated squamous cell carcinoma to develop novel therapeutic interventions

Andrew C. White and William E. Lowry
Pages 318 - 321
http://dx.doi.org/10.4161/sgtp.18088
Abstract | Full Text | PDF

The small GTPase Ras is activated in a high proportion of human cancers. Attempts to clinically block Ras activity through pharmacological means has proven largely ineffective thus far. We employed an inducible mouse model of squamous cell carcinoma (SCC) to study the effect of Ras activation and show that hair follicle stem cells (HFSCs) are a cell of origin for SCC, whereas their more restricted progeny cannot serve as cancer cells of origin and are refractory to Ras activation. We propose that by identifying the unique mechanisms by which HFSCs are mobilized to initiate Ras mediated tumorigenesis, the molecular process behind SCC can be more completely elucidated and context dependent activities for Ras more clearly defined. Here, we summarize our recent results and point to future experiments designed to create novel therapeutics by exploiting the differential sensitivities of various cells within the epidermis to Ras activation.

Open Access Article

Polarization of migrating cortical neurons by Rap1 and N-cadherin: Revisiting the model for the Reelin signaling pathway

Yves Jossin
Pages 322 - 328
http://dx.doi.org/10.4161/sgtp.18283
Abstract | Full Text | PDF

Neuronal migration is essential for the development of the cerebral cortex. Mutations leading to defective migration are associated with numerous brain pathologies. An important challenge in the field is to understand the intrinsic and extrinsic mechanisms that regulate neuronal migration during normal development and in disease. Many small GTPases are expressed in the central nervous system during embryonic development. Recent findings have shown that Rap1 and its downstream partners Ral, Rac and Cdc42 are involved in the maintenance of N-Cadherin at the plasma membrane which is necessary for the correct polarization of migrating neurons. The activation of Rap1 is triggered by Reelin, an extracellular protein known for its role in the organization of the cortex into layers of neurons. In the absence of Reelin, neurons exhibit a broader and irregular pattern of positioning. The prevailing model suggests that Reelin signals to neurons during the last step of their migration, a notion that is inconsistent with new data describing an effect of Reelin on early steps of migration. In regard to these recent findings I suggest a revised model, which I call the “polarity model,” that further refines our understanding of the developmental function played by Reelin and its downstream small GTPases.

Open Access Article

Breaking up is hard to do: RalA, mitochondrial fission and cancer

David F. Kashatus and Christopher M. Counter
Pages 329 - 333
http://dx.doi.org/10.4161/sgtp.18284
Abstract | Full Text | PDF

The small GTPases RalA and RalB are activated downstream of oncogenic Ras. While activation of RalA is critically important for tumor initiation and growth of Ras-driven cancers, the highly similar small GTPase RalB is implicated in cell survival and metastasis. This difference in function between these two related proteins maps to the C-terminus, a 30 amino acid region that regulates subcellular localization and contains several potential phosphorylation sites. Here we discuss our recent evidence that phosphorylation by the mitotic kinase Aurora A promotes RalA relocalization to mitochondrial membranes, where it recruits the effector RalBP1 and the large dynamin-related GTPase Drp1 to promote mitochondrial fission. As upregulation of both RalA and Aurora A have been observed in human tumors, and phosphorylation of RalA at the site targeted by Aurora A promotes tumorigenesis, it is possible that regulation of mitochondrial fission is one mechanism by which RalA promotes cancer.