Rnd Proteins Research Articles

Rapostlin Is a Novel Effector of Rnd2 GTPase Inducing Neurite Branching

Rho family GTPases are central regulators of neuronal morphology. Recently, Rnd proteins, Rnd1, Rnd2, and Rnd3/RhoE, have been identified as new members of Rho family GTPases. Of these, Rnd2 is specifically expressed in neurons in brain; however, the signaling pathways of Rnd2 are not known. Here we have performed a yeast two-hybrid screen using Rnd2 as a bait and identified a novel Rnd2-effector protein, expressed predominantly in brain. We named it Rapostlin (apostle of Rnd2). Rapostlin has two functional domains, Fer-CIP4 homology (FCH) domain at the amino terminus and SH3 (Src homology 3) domain at the carboxyl terminus. In in vitro binding assays, Rapostlin specifically binds to Rnd2 among the Rho family GTPases in a GTP-dependent manner, and the Rnd2-binding domain of Rapostlin is localized between FCH and SH3 domains. Rapostlin directly binds to microtubules, and the amino-terminal region containing the FCH domain of Rapostlin is essential for this interaction. In PC12 cells, Rapostlin induces neurite branching in response to Rnd2, and at least the amino-terminal region of Rapostlin is necessary for this activity. Therefore, Rapostlin is the first effector of Rnd2, regulating neurite branch formation.

Dispatching the Hedgehog Signal

The mammalian sonic hedgehog (Shh) protein functions as a secreted ligand that has critical roles in regulation of development. Hedgehog (Hh) proteins are modified by addition of two lipid moieties, a cholesteryl moiety and palmitate, which might be expected to promote association of the Shh protein with membranes of the cells that produce it. However, developmental signaling through Shh requires long-range interactions with receptors on cells that are many cells distant from the cell that produces Shh. In Drosophila , genetic evidence indicates that the protein Dispatched (Disp) takes part in presentation or release of the Hh protein. Ma et al. characterized the mouse homolog of Disp called mDispA. Disruption of the gene encoding mDispA caused developmental defects that reflected a lack of long-range signaling by Shh. Experiments with cultured embryonic fibroblasts lacking mDispA showed that release and extracellular accumulation of Shh were blocked. Exactly how mDispA influences release of Shh remains unclear. However, the Disp proteins are similar in structure to prokaryotic proteins called RND permeases that function as transporters to mediate efflux of molecules from cells. Similarity to RND proteins is also shared by Patched, a protein that functions in responses to Hh at target cells. Thus, the hedgehog pathway appears to have two participants from the RND family that have evolved to serve distinct roles in release and response to Hh ligands. Y. Ma, A. Erkner, R. Gong, S. Yao, J. Taipale, K. Basler, P. A. Beachy, Hedgehog-mediated patterning of the mammalian embryo requires transporter-like function of dispatched. Cell 111 , 63-75 (2002). [Online Journal]

Crystal structure of Rnd3/RhoE: functional implications

The Rnd proteins constitute an exceptional subfamily within the Rho GTPase family. They possess extended chains at both termini and four prominent amino acid deviations causing GTPase deficiency. Herein, we report the crystal structure of the Rnd3/RhoE G-domain (amino acids 19–200) at 2.0 Å resolution. This is the first GTP-structure of a Rho family member which reveals a similar fold but striking differences from RhoA concerning (i) GTPase center, (ii) charge distribution at several surface areas, (iii) C3-transferase binding site and (iv) interacting interfaces towards RhoA regulators and effectors.

Socius is a novel Rnd GTPase-interacting protein involved in disassembly of actin stress fibers.

Rho family small GTPases are key regulators of the actin cytoskeleton in various cell types. The Rnd proteins, Rnd1, Rnd2, and Rnd3/RhoE, have been recently identified as new members of the Rho family of GTPases, and expression of Rnd1 or Rnd3 in fibroblasts causes the disassembly of actin stress fibers and the retraction of the cell body to produce extensively branching cellular processes. Here we have performed a yeast two-hybrid screening by using Rnd1 as bait and identified a novel protein that specifically binds to Rnd GTPases. We named this protein Socius. Socius directly binds to Rnd GTPases through its COOH-terminal region. When transfected into COS-7 cells, Socius is translocated to the cell periphery in response to Rnd1 and Rnd3 and colocalized with the GTPases. While expression of wild-type Socius in Swiss 3T3 fibroblasts has little effect on the actin cytoskeleton, the expression of a membrane-targeted form of Socius, containing a COOH-terminal farnesylation motif (Socius-CAAX), induces a dramatic loss of stress fibers. The inhibitory effect of Socius-CAAX on stress fiber formation is enhanced by truncation of its NH(2) terminus. On the other hand, the expression of Socius-CAAX or its NH(2) terminus-truncated form suppresses the Rnd-induced retraction of the cell body and the production of extensively branching cellular processes, although the disassembly of stress fibers is observed. We propose that Socius participates in the Rnd GTPase-induced signal transduction pathways, leading to reorganization of the actin cytoskeleton.

Rnd proteins: a new family of Rho-related proteins that interfere with the assembly of filamentous actin structures and cell adhesion.

Rho family proteins control actin organization. The Rnd proteins form a distinct branch of the Rho family: Rnd1 is expressed mostly in brain and liver, Rnd2 is highly expressed in testis and Rnd3 shows a ubiquitous, very low expression. In brain, Rnd1 is found in specialized neurons, mainly in the cortex, hippocampus and substantia nigra. Rnd1 exchanges GTP rapidly, has a low affinity for GDP, and lacks intrinsic GTPase activity, suggesting that in the cell it is constitutively in the GTP-bound form. Expression of Rnd1 in Swiss 3T3 fibroblasts inhibits the formation of actin stress fibers and induces a loss of focal adhesions and cell/cell contacts, leading to cell rounding (hence Rnd for “round”). In Swiss 3T3 fibroblasts and MDCK cells, Rnd1 localizes to adherens junctions. Thus, Rnd proteins are involved in rearrangements of the actin cytoskeleton and changes in cell adhesion that might play essential roles in cell migration and tumor invasion.KeywordsStress FiberActin Stress FiberPhosphorylate Myosin Light ChainStress Fiber FormationMyosin Light Chain PhosphataseThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.