c-Abl, Lamellipodin, and Ena/VASP Proteins Cooperate in Dorsal Ruffling of Fibroblasts and Axonal Morphogenesis

Randall Division of Cell and Molecular Biophysics, King's College London, New Hunt's House, Guy's Campus, London SE1 1UL, UK.
Current biology: CB (Impact Factor: 9.57). 05/2010; 20(9):783-91. DOI: 10.1016/j.cub.2010.03.048
Source: PubMed


Tight regulation of cell motility is essential for many physiological processes, such as formation of a functional nervous system and wound healing. Drosophila Abl negatively regulates the actin cytoskeleton effector protein Ena during neuronal development in flies, and it has been postulated that this may occur through an unknown intermediary. Lamellipodin (Lpd) regulates cell motility and recruits Ena/VASP proteins (Ena, Mena, VASP, EVL) to the leading edge of cells. However, the regulation of this recruitment has remained unsolved.
Here we show that Lpd is a substrate of Abl kinases and binds to the Abl SH2 domain. Phosphorylation of Lpd positively regulates the interaction between Lpd and Ena/VASP proteins. Consistently, efficient recruitment of Mena and EVL to Lpd at the leading edge requires Abl kinases. Furthermore, transient Lpd phosphorylation by Abl kinases upon netrin-1 stimulation of primary cortical neurons positively correlates with an increase in Lpd-Mena coprecipitation. Lpd is also transiently phosphorylated by Abl kinases upon platelet-derived growth factor (PDGF) stimulation, regulates PDGF-induced dorsal ruffling of fibroblasts and axonal morphogenesis, and cooperates with c-Abl in an Ena/VASP-dependent manner.
Our findings suggest that Abl kinases positively regulate Lpd-Ena/VASP interaction, Ena/VASP recruitment to Lpd at the leading edge, and Lpd-Ena/VASP function in axonal morphogenesis and in PDGF-induced dorsal ruffling. Our data do not support the suggested negative regulatory role of Abl for Ena. Instead, we propose that Lpd is the hitherto unknown intermediary between Abl and Ena/VASP proteins.

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Available from: Matthias Krause, Jul 20, 2014
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    • "It will be interesting to investigate in future studies whether our observations of filopodia overgrowth in Abl neurons also involves dysregulation of Dia-related formins. In vertebrates, c-Abl and Ena/VASP also regulate the leading edge mediated by a specific intermediary protein, Lamellipodin, which is required for lamellipodia formation during axonal morphogenesis [53], [54]. It will be important to identify the Drosophila homologous gene(s) encoding Lamellipodin-related protein(s) in order to elucidate the mechanism of enhancing muscle lamellipodia formation and its relation to muscle cell movement and fusion, and to explain the apparently opposite effects on neuronal growth. "
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    ABSTRACT: Abelson tyrosine kinase (Abl) is known to regulate axon guidance, muscle development, and cell-cell interaction in vivo. The Drosophila primary culture system offers advantages in exploring the cellular mechanisms mediated by Abl with utilizing various experimental manipulations. Here we demonstrate that single-embryo cultures exhibit stage-dependent characteristics of cellular differentiation and developmental progression in neurons and myocytes, as well as nerve-muscle contacts. In particular, muscle development critically depends on the stage of dissociated embryos. In wild-type (WT) cultures derived from embryos before stage 12, muscle cells remained within cell clusters and were rarely detected. Interestingly, abundant myocytes were spotted in Abl mutant cultures, exhibiting enhanced myocyte movement and fusion, as well as neuron-muscle contacts even in cultures dissociated from younger, stage 10 embryos. Notably, Abl myocytes frequently displayed well-expanded lamellipodia. Conversely, Abl neurons were characterized with fewer large veil-like lamellipodia, but instead had increased numbers of filopodia and darker nodes along neurites. These distinct phenotypes were equally evident in both homo- and hetero-zygous cultures (Abl/Abl vs. Abl/+) of different alleles (Abl(1) and Abl(4) ) indicating dominant mutational effects. Strikingly, in WT cultures derived from stage 10 embryos, high temperature (HT) incubation promoted muscle migration and fusion, partially mimicking the advanced muscle development typical of Abl cultures. However, HT enhanced neuronal growth with increased numbers of enlarged lamellipodia, distinct from the characteristic Abl neuronal morphology. Intriguingly, HT incubation also promoted Abl lamellipodia expansion, with a much greater effect on nerve cells than muscle. Our results suggest that Abl is an essential regulator for myocyte and neuron development and that high-temperature incubation partially mimics the faster muscle development typical of Abl cultures. Despite the extensive alterations by Abl mutations, we observed myocyte fusion events and nerve-muscle contact formation between WT and Abl cells in mixed WT and Abl cultures derived from labeled embryos.
    PLoS ONE 01/2014; 9(1):e86438. DOI:10.1371/journal.pone.0086438 · 3.23 Impact Factor
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    • "As the levels of Mena should be increased to accelerate wound healing [18, 28], lithium-induced β-catenin increases Mena transcription in these cases [26]. "
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    ABSTRACT: The Ena/VASP (enabled/vasodilator stimulated phosphoprotein) family includes the binding actin proteins such as mammalian Ena (Mena), VASP, and Ena-VASP-like. It is known that the perturbation of actin cycle could determine alteration in the mobility of cells and in consequence of organogenesis. Few recent studies have revealed that Mena protein could play a role in breast or pancreatic carcinogenesis. Based on our researches, we observed that the intensity of Mena expression increased from premalignant to malignant lesions in some organs such as large bowel, stomach, cervix, and salivary glands. These findings prove that Mena could be a marker of premalignant epithelial lesions. In premalignant lesions, it could be helpful to define more accurately the risk for malignant transformation. In malignant tumors, correlation of expression of its splice variants could indicate metastatic behavior. In conclusion, we consider that it is necessary to analyze the expression of Mena splice variants in a higher number of cases, in different epithelial lesions, and also in experimental studies to define its exact role in carcinogenesis and also its possible prognostic and predictive values.
    07/2013; 2013(3):365192. DOI:10.1155/2013/365192
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    • "The Caenorhabditis elegans Lpd ortholog MIG-10 has important roles in neuronal migration, polarization, and axon guidance (Adler et al., 2006; Chang et al. 2006; Quinn et al. 2006). Recent studies in mammals have shown that knockdown of Lpd decreases the length of axons (Michael et al., 2010). It has also been reported that Lpd regulates migration of pyramidal neurons through the regulation of serum response factor (SRF) transcription factor activity (Pinheiro et al., 2011). "
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    ABSTRACT: Semaphorins have been identified as repulsive guidance molecules in the developing nervous system. We recently reported that the semaphorin 4D (Sema4D) receptor Plexin-B1 induces repulsion in axon and dendrites by functioning as a GTPase-activating protein (GAP) for R-Ras and M-Ras, respectively. In axons, Sema4D stimulation induces growth cone collapse, and downregulation of R-Ras activity by Plexin-B1-mediated GAP activity is required for the action. Axonal R-Ras GAP activity downregulates phosphatidylinositol 3-kinase signaling pathway, and thereby induces inactivation of a microtubule assembly promoter protein, CRMP-2. However, in contrast to the well studied roles of semaphorins and plexins in axonal guidance, signaling molecules linking M-Ras GAP to dendritic cytoskeleton remain obscure. Here we identified an Ena/VASP ligand, Lamellipodin (Lpd), as a novel effector of M-Ras in dendrites. Lpd was expressed in F-actin-rich distal dendritic processes and was required for both basal and M-Ras-mediated dendrite development. Subcellular fractionation showed M-Ras-dependent membrane translocation of Lpd, which was suppressed by Sema4D. Furthermore, the Ena/VASP-binding region within Lpd was required for dendrite development, and its membrane targeting was sufficient to overcome the Sema4D-mediated reduction of dendritic outgrowth and disappearance of F-actin from distal dendrites. Furthermore, in utero electroporation experiments also indicated that regulation of the M-Ras-Lpd system by the GAP activity of Plexin is involved in the normal development of cortical dendrites in vivo. Overall, our study sheds light on how repulsive guidance molecules regulate actin cytoskeleton in dendrites, revealing a novel mechanism that the M-Ras-Lpd system regulates actin-based dendrite remodeling by Sema/Plexin in rats or mice of either sex.
    The Journal of Neuroscience : The Official Journal of the Society for Neuroscience 06/2012; 32(24):8293-305. DOI:10.1523/JNEUROSCI.0799-12.2012 · 6.34 Impact Factor
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