Phosphorylation of the Zebrafish M6Ab at Serine 263 Contributes to Filopodium Formation in PC12 Cells and Neurite Outgrowth in Zebrafish Embryos

Huang, Kai; Chen, Gen Der; Cheng, Chia Hsiung; Liao, Kuan Ya; Hung, Ching‐Wen; Chang, Geen Dong; Hwang, Pung Pung; Lin, Shu Yu; Tsai, Ming Chieh; Khoo, Kay‐Hooi; Lee, Ming Ting; Huang, Chang Jen

doi:10.1371/journal.pone.0026461

Cited by 14 publications

(14 citation statements)

References 38 publications

(62 reference statements)

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“…The phosphorylation state of M6a at Y251 seems not to be involved in the formation of filopodia-like processes in hippocampal neurons. In agreement with these findings, phosphorylation at Ser263 of the M6azebrafish ortholog M6Ab promotes filopodia formation, but not neurite extension, in PC12 cells (Huang et al, 2011). Moreover, M6a lacking intracellular phosphorylation sites for casein-kinase 2, and protein PKC-expressing neurons bore filopodia, but the motility of the protrusions was slow compared with that of M6a wt .…”

Section: Discussionsupporting

confidence: 74%

“…Accordingly, we found that the structure of EC2 is critical to induce filopodium outgrowth and synaptogenesis (Fuchsova et al, ) and that mutations in M6a intracellular sites, which are motifs for protein kinase C (PKC) and casein kinase‐2 phosphorylation, decrease the motility of filopodia induced by M6a (Brocco et al, ). The zebrafish M6a ortholog, which lacks a phosphorylation site for PKC, fails to induce neurite extension in PC12 cells (Huang et al, ). Also, M6a depends on membrane lipid microdomain association and requires the activity of Src kinases and mitogen‐activated protein kinases (MAPK) for its filopodium induction (Scorticati et al, ).…”

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confidence: 99%

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Tyrosine 251 at the C‐terminus of neuronal glycoprotein M6a is critical for neurite outgrowth

Formoso

Billi

Frasch

et al. 2014

J of Neuroscience Research

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Neuronal glycoprotein M6a is involved in neuronal plasticity, promoting neurite and filopodia outgrowth and, likely, synaptogenesis. Polymorphisms in the human M6a gene GPM6A have recently been associated with mental illnesses such as schizophrenia, bipolar disorders, and claustrophobia. Nevertheless, the molecular bases underlying these observations remain unknown. We have previously documented that, to induce filopodia formation, M6a depends on the association of membrane lipid microdomains and the activation of Src and mitogen-activated protein kinase kinases. Here, in silico analysis of the phosphorylation of tyrosine 251 (Y251) at the C-terminus of M6a showed that it could be a target of Src kinases. We examined whether phosphorylation of M6a at Y251 affects neurite and filopodia outgrowth and the targets involved in its signal propagation. This work provides evidence that the Src kinase family and the phosphatidylinositide 3-kinase (PI3K), but not Ras, participate in M6a signal cascade leading to neurite/filopodia outgrowth in hippocampal neurons and murine neuroblastoma N2a cells. Phosphorylation of M6a at Y251 is essential only for neurite outgrowth by the PI3K/AKT-mediated pathway and, moreover, rescues the inhibition caused by selective Src inhibitor and external M6a monoclonal antibody treatment. Thus, we suggest that phosphorylation of M6a at Y251 is critical for a specific stage of neuronal development and triggers redundant signaling pathways leading to neurite extension.

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Section: Discussionsupporting

confidence: 74%

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confidence: 99%

Tyrosine 251 at the C‐terminus of neuronal glycoprotein M6a is critical for neurite outgrowth

Formoso

Billi

Frasch

et al. 2014

J of Neuroscience Research

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“…M6a is a membrane glycoprotein encoded by the gene GPM6A. M6a, together with M6b, proteolipid protein (PLP) and DM20, belongs to the PLP family, and has been related to neuronal plasticity in different experimental models (Lagenaur et al 1992;Alfonso et al 2005;Michibata et al 2009;Huang et al 2011;Sato et al 2011a;Zappia et al 2012). M6a positively contributes to neuritogenesis, filopodia outgrowth and synapse formation in M6a-over-expressing neurons, but its mechanism of action remains unknown (Alfonso et al 2005;Fuchsova et al 2009;Formoso et al 2015).…”

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confidence: 99%

Filopodia formation driven by membrane glycoprotein M6a depends on the interaction of its transmembrane domains

Formoso

Garcia

Frasch

et al. 2015

Journal of Neurochemistry

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Membrane glycoprotein M6a, which belongs to the tetraspan proteolipid protein family, promotes structural plasticity in neurons and cell lines by unknown mechanisms. This glycoprotein is encoded by Gpm6a, a stress-regulated gene. The hippocampus of animals chronically stressed by either psychosocial or physical stressors shows decreased M6a expression. Stressed Gpm6a-null mice develop a claustrophobia-like phenotype. In humans, de novo duplication of GPM6A results in learning/behavioral abnormalities, and two single-nucleotide polymorphisms (SNPs) in the non-coding region are linked to mood disorders. Here, we studied M6a dimerization in neuronal membranes and its functional relevance. We showed that the self-interaction of M6a transmembrane domains (TMDs) might be driving M6a dimerization, which is required to induce filopodia formation. Glycine mutants located in TMD2 and TMD4 of M6a affected its dimerization, thus preventing M6a-induced filopodia formation in neurons. In silico analysis of three non-synonymous SNPs located in the coding region of TMDs suggested that these mutations induce protein instability. Indeed, these SNPs prevented M6a from being functional in neurons, owing to decreased stability, dimerization or improper folding. Interestingly, SNP3 (W141R), which caused endoplasmic reticulum retention, is equivalent to that mutated in PLP1, W161L, which causes demyelinating Pelizaeus-Merzbacher disease.

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“…The 293T cells were maintained at 37 °C in minimal essential medium-alpha medium, 10 % fetal bovine serum, 100 mg/ml streptomycin, and 100 U/ml penicillin. Plasmid transfection and cell collection were done with standard procedures as described previously [67]. Transfected 293T cells collected from one 6-well plate were lysed in 0.1 ml immunoprecipitation lysis buffer (150 mM NaCl, 20 mM HEPES (pH 7.2), 10 mM NaF, 1 mM EDTA, 0.5 % NP-40, 1 mM Na3VO4, 1 mM PMSF, and 1 mM DTT), then sonicated for 10 s three times using a UP50H machine at an 80 % power level (Dr. Hielscher, Teltow, Germany).…”

Section: Methodsmentioning

confidence: 99%

Protein tyrosine phosphatase receptor type O (Ptpro) regulates cerebellar formation during zebrafish development through modulating Fgf signaling

Liao

Cheng

Hung

et al. 2013

Cell. Mol. Life Sci.

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Protein activities controlled by receptor protein tyrosine phosphatases (RPTPs) play comparably important roles in transducing cell surface signals into the cytoplasm by protein tyrosine kinases. Previous studies showed that several RPTPs are involved in neuronal generation, migration, and axon guidance in Drosophila, and the vertebrate hippocampus, retina, and developing limbs. However, whether the protein tyrosine phosphatase type O (ptpro), one kind of RPTP, participates in regulating vertebrate brain development is largely unknown. We isolated the zebrafish ptpro gene and found that its transcripts are primarily expressed in the embryonic and adult central nervous system. Depletion of zebrafish embryonic Ptpro by antisense morpholino oligonucleotide knockdown resulted in prominent defects in the forebrain and cerebellum, and the injected larvae died on the 4th day post-fertilization (dpf). We further investigated the function of ptpro in cerebellar development and found that the expression of ephrin-A5b (efnA5b), a Fgf signaling induced cerebellum patterning factor, was decreased while the expression of dusp6, a negative-feedback gene of Fgf signaling in the midbrain-hindbrain boundary region, was notably induced in ptpro morphants. Further analyses demonstrated that cerebellar defects of ptpro morphants were partially rescued by inhibiting Fgf signaling. Moreover, Ptpro physically interacted with the Fgf receptor 1a (Fgfr1a) and dephosphorylated Fgfr1a in a dose-dependant manner. Therefore, our findings demonstrate that Ptpro activity is required for patterning the zebrafish embryonic brain. Specifically, Ptpro regulates cerebellar formation during zebrafish development through modulating Fgf signaling.Electronic supplementary materialThe online version of this article (doi:10.1007/s00018-013-1259-7) contains supplementary material, which is available to authorized users.

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Phosphorylation of the Zebrafish M6Ab at Serine 263 Contributes to Filopodium Formation in PC12 Cells and Neurite Outgrowth in Zebrafish Embryos

Cited by 14 publications

References 38 publications

Tyrosine 251 at the C‐terminus of neuronal glycoprotein M6a is critical for neurite outgrowth

Tyrosine 251 at the C‐terminus of neuronal glycoprotein M6a is critical for neurite outgrowth

Filopodia formation driven by membrane glycoprotein M6a depends on the interaction of its transmembrane domains

Protein tyrosine phosphatase receptor type O (Ptpro) regulates cerebellar formation during zebrafish development through modulating Fgf signaling

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