Myosin-Va-interacting protein, RILPL2, controls cell shape and neuronal morphogenesis via Rac signaling
ABSTRACT Neuronal morphology plays an essential role in neuronal function. The establishment and maintenance of neuronal morphology is intimately linked to the actin cytoskeleton; however, the molecular mechanisms that regulate changes in neuronal morphology are poorly understood. Here we identify a novel myosin-Va (MyoVa)-interacting protein, RILPL2, which regulates cellular morphology. Overexpression of this protein in young or mature hippocampal neurons results in an increase in the number of spine-like protrusions. By contrast, knockdown of endogenous RILPL2 in neurons by short hairpin RNA (shRNA) interference results in reduced spine-like protrusions, a phenotype rescued by overexpression of an shRNA-insensitive RILPL2 mutant, suggesting a role for RILPL2 in both the establishment and maintenance of dendritic spines. Interestingly, we demonstrate that RILPL2 and the Rho GTPase Rac1 form a complex, and that RILPL2 is able to induce activation of Rac1 and its target, p21-activated kinase (Pak). Notably, both RILPL2-mediated morphological changes and activation of Rac1-Pak signaling were blocked by expression of a truncated tail form of MyoVa or MyoVa shRNA, demonstrating that MyoVa is crucial for proper RILPL2 function. This might represent a novel mechanism linking RILPL2, the motor protein MyoVa and Rac1 with neuronal structure and function.
- SourceAvailable from: Nicholas Thomas Hertz
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- "Myosin Va is also implicated in transporting endoplasmic reticulum in Purkinje neuron dendritic spines (Wagner et al., 2011). Myosin Va knockdown in neuronal cultures drastically reduces spine density (Lisé et al., 2009). A recent examination of the Myosin Va neurological mutant mice flailer, which express a dominant-negative Myosin Va fusion protein lacking the actin binding domain, revealed a reduction of mature dendritic spines and an increase in dendritic filopodia (Yoshii et al., 2013). "
ABSTRACT: Mammalian Sterile 20 (Ste20)-like kinase 3 (MST3) is a ubiquitously expressed kinase capable of enhancing axon outgrowth. Whether and how MST3 kinase signaling might regulate development of dendritic filopodia and spine synapses is unknown. Through shRNA-mediated depletion of MST3 and kinase-dead MST3 expression in developing hippocampal cultures, we found that MST3 is necessary for proper filopodia, dendritic spine, and excitatory synapse development. Knockdown of MST3 in layer 2/3 pyramidal neurons via in utero electroporation also reduced spine density in vivo. Using chemical genetics, we discovered thirteen candidate MST3 substrates and identified the phosphorylation sites. Among the identified MST3 substrates, TAO kinases regulate dendritic filopodia and spine development, similar to MST3. Furthermore, using stable isotope labeling by amino acids in culture (SILAC), we show that phosphorylated TAO1/2 associates with Myosin Va and is necessary for its dendritic localization, thus revealing a mechanism for excitatory synapse development in the mammalian CNS. Copyright © 2014 Elsevier Inc. All rights reserved.Neuron 12/2014; 84(5):968-82. DOI:10.1016/j.neuron.2014.10.025 · 15.98 Impact Factor
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- "Sequence alignment was used to divide Rilpl1 at positions equivalent to those in Rilpl2 (ΔC, aa 1–288; ΔN, aa 289–406). The ΔC construct of both proteins contains the RH1 domain and in Rilpl2 the MyoVa interaction domain (Lisé et al., 2009). The ΔN construct of both proteins contains the RH2 domain and the Rab36 interaction domain (Figure 1A; Matsui et al., 2012). "
ABSTRACT: The primary cilium is a microtubule-based structure found in most cell types in mammals. Disruption of cilium function causes a diverse set of human diseases collectively known as ciliopathies. We report that Rab effector-related proteins Rilpl1 and Rilpl2 regulate protein localization in the primary cilium. Rilpl2 was initially identified as up-regulated in ciliating mouse tracheal epithelial cells. Rilpl1 and Rilpl2 both localize to the primary cilium and centrosome, Rilpl1 specifically to the distal end of the mother centriole. Live cell microscopy reveals that Rilpl2 primary cilium localization is dynamic and that it is associated with tubulovesicular structures at the base of the cilium. Depletion of Rilpl1 and Rilpl2 results in accumulation of signaling proteins in the ciliary membrane and prevents proper epithelial cell organization in three-dimensional culture. These data suggest that Rilp-like proteins function in regulation of ciliary membrane protein concentration by promoting protein removal from the primary cilium.Molecular biology of the cell 12/2012; 24(4). DOI:10.1091/mbc.E12-08-0598 · 5.98 Impact Factor