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Inverse Expression Levels of EphrinA3 and EphrinA5 Contribute to Dopaminergic Differentiation of Human SH-SY5Y Cells

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Abstract

Two key principles underlying successful cellular therapies for Parkinson’s disease (PD) are appropriate differentiation of dopaminergic (DA) neurons from transplanted cells and precise axon growth. EphrinAs, a subclass of ephrins, act as axon guidance molecules and are highly expressed in DA brain regions. Existing evidences indicate that they act as either repulsion or attraction signals to guide axon growth. This study investigated whether ephrinAs are involved in DA neuron differentiation. Data from miRCURY™ LNA mRNAs/microRNAs microarrays and quantitative real-time polymerase chain reaction (qRT-PCR) showed upregulated ephrinA3 mRNA (EFNA3) and downregulated ephrinA5 mRNA (EFNA5) during DA neuron differentiation. In addition, hsa-miR-4271 was downregulated, which could influence EFNA3 translation. Furthermore, immunofluorescence (IF) and western blotting confirmed the mRNA results and showed increased ephrinA3 and decreased ephrinA5 protein levels in differentiating DA neurons. Taken together, our results indicate that inverse expression levels of ephrinA3 and ephrinA5, which are possibly influenced by microRNAs, contribute to DA neuron differentiation by guiding axon growth.

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References

  • Aschrafi A, Schwechter AD, Mameza MG, Natera-Naranjo O, Gioio AE, Kaplan BB (2008) MicroRNA-338 regulates local cytochrome c oxidase IV mRNA levels and oxidative phosphorylation in the axons of sympathetic neurons. J Neurosci 28:12581–12590

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bartel DP (2009) MicroRNAs target recognition and regulatory functions. Cell 136:215–233

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Bové J, Prou D, Perier C, Przedborski S (2005) Toxin-induced models of Parkinson’s disease. NeuroRx 2:484–494

    Article  PubMed  PubMed Central  Google Scholar 

  • Brownlee H, Gao PP, Frisén J, Dreyfus C, Zhou R, Black IB (2000) Multiple ephrins regulate hippocampal neurite outgrowth. J Comp Neurol 425:315–322

    Article  CAS  PubMed  Google Scholar 

  • Bu L, Li R, Liu H, et al. (2014) Intrastriatal transplantation of retinal pigment epithelial cells for the treatment of Parkinson disease: in vivo longitudinal molecular imaging with 18F-P3BZA PET/CT. Radiology 272:174–183

    Article  PubMed  PubMed Central  Google Scholar 

  • Bush JO, Soriano P (2012) Eph/ephrin signaling: genetic, phosphoproteomic, and transcriptomic approaches. Semin Cell Dev Biol 23:26–34

    Article  CAS  PubMed  Google Scholar 

  • Cang J, Wang L, Stryker MP, Feldheim DA (2008) Roles of ephrin-as and structured activity in the development of functional maps in the superior colliculus. J Neurosci 28:11015–11023

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Carmona MA, Murai KK, Wang L, Roberts AJ, Pasquale EB (2009) Glial ephrin-A3 regulates hippocampal dendritic spine morphology and glutamate transport. Proc Natl Acad Sci U S A 106:12524–12529

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Cheng H-J, Nakamoto M, Bergemann AD, Flanagan JG (1995) Complementary gradients in expression and binding of ELF-1 and Mek4 in development of the topographic retinotectal projection map. Cell 82:371–381

    Article  CAS  PubMed  Google Scholar 

  • Cowan CA, Henkemeyer M (2002) Ephrins in reverse, park and drive. Trends Cell Biol 12:339–346

    Article  CAS  PubMed  Google Scholar 

  • Cutforth T, Moring L, Mendelsohn M, et al. (2003) Axonal ephrin-as and odorant receptors: coordinate determination of the olfactory sensory map. Cell 114:311–322

    Article  CAS  PubMed  Google Scholar 

  • Dajas-Bailador F, Bonev B, Garcez P, Stanley P, Guillemot F, Papalopulu N (2012) microRNA-9 regulates axon extension and branching by targeting Map1b in mouse cortical neurons. Nat Neurosci 15:697–699

    Article  CAS  Google Scholar 

  • Davy A, Robbins SM (2000) Ephrin-A5 modulates cell adhesion and morphology in an integrin-dependent manner. EMBO J 19:5396–5405

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Davy A, Soriano P (2005) Ephrin signaling in vivo: look both ways. Dev Dyn 232:1–10

    Article  CAS  PubMed  Google Scholar 

  • Deumens R, Blokland A, Prickaerts J (2002) Modeling Parkinson’s disease in rats: an evaluation of 6-OHDA lesions of the nigrostriatal pathway. Exp Neurol 175:303–317

    Article  CAS  PubMed  Google Scholar 

  • Drescher U, Kremoser C, Handwerker C, Loschinger J, Noda M, Bonhoeffer F (1995) In vitro guidance of retinal ganglion cell axons by RAGS, a 25 kDa tectal protein related to ligands for Eph receptor tyrosine kinases. Cell 82:359–370

    Article  CAS  PubMed  Google Scholar 

  • Dufour A, Seibt J, Passante L, et al. (2003) Area specificity and topography of thalamocortical projections are controlled by ephrin/Eph genes. Neuron 39:453–465

    Article  CAS  PubMed  Google Scholar 

  • Feldheim DA, Kim YI, Bergemann AD, Frisén J, Barbacid M, Flanagan JG (2000) Genetic analysis of ephrin-A2 and ephrin-A5 shows their requirement in multiple aspects of retinocollicular mapping. Neuron 25:563–574

    Article  CAS  PubMed  Google Scholar 

  • Flanagan JG (2006) Neural map specification by gradients. Curr Opin Neurobiol 16:59–66

    Article  CAS  PubMed  Google Scholar 

  • Flanagan JG, Vanderhaeghen P (1998) The ephrins and Eph receptors in neural development. Annu Rev Neurosci 21:309–345

    Article  CAS  PubMed  Google Scholar 

  • Frisén J, Yates PA, McLaughlin T, Friedman GC, O’Leary DD, Barbacid M (1998) Ephrin-A5 (AL-1/RAGS) is essential for proper retinal axon guidance and topographic mapping in the mammalian visual system. Neuron 20:235–243

    Article  PubMed  Google Scholar 

  • Grealish S, Xie L, Kelly M, Dowd E (2008) Unilateral axonal or terminal injection of 6-hydroxydopamine causes rapid-onset nigrostriatal degeneration and contralateral motor impairments in the rat. Brain Res Bull 77:312–319

    Article  CAS  PubMed  Google Scholar 

  • Grunwald IC, Korte M, Adelmann G, et al. (2004) Hippocampal plasticity requires postsynaptic ephrinBs. Nat Neurosci 7:33–40

    Article  CAS  PubMed  Google Scholar 

  • Hattori M, Osterfield M, Flanagan JG (2000) Regulated cleavage of a contact-mediated axon repellent. Science 289:1360–1365

    Article  CAS  PubMed  Google Scholar 

  • Hengst U, Cox LJ, Macosko EZ, Jaffrey SR (2006) Functional and selective RNA interference in developing axons and growth cones. J Neurosci 26:5727–5732

    Article  CAS  PubMed  Google Scholar 

  • Heuer A, Lelos MJ, Kelly CM, Torres EM, Dunnett SB (2013) Dopamine-rich grafts alleviate deficits in contralateral response space induced by extensive dopamine depletion in rats. Exp Neurol 247:485–495

    Article  CAS  PubMed  Google Scholar 

  • Himanen JP, Rajashankar KR, Lackmann M, Cowan CA, Henkemeyer M, Nikolov DB (2001) Crystal structure of an Eph receptor-ephrin complex. Nature 414:933–938

    Article  CAS  PubMed  Google Scholar 

  • Holmberg J, Frisén J (2002) Ephrins are not only unattractive. Trends Neurosci 25:239–243

    Article  CAS  PubMed  Google Scholar 

  • Holmberg J, Clarke DL, Frisén J (2000) Regulation of repulsion versus adhesion by different splice forms of an Eph receptor. Nature 408:203–206

    Article  CAS  PubMed  Google Scholar 

  • Kiessling F (2014) Science to practice: cellular therapy of Parkinson disease—a new radiotracer to target transplanted dopaminergic cells with PET. Radiology 272:1–3

    Article  PubMed  Google Scholar 

  • Kim VN (2005) MicroRNA biogenesis: coordinated cropping and dicing. Nat Rev Mol Cell Biol 6:376–385

    Article  CAS  PubMed  Google Scholar 

  • Knoll B, Drescher U (2002) Ephrin-as as receptors in topographic projections. Trends Neurosci 25:145–149

    Article  CAS  PubMed  Google Scholar 

  • Knoll B, Zarbalis K, Wurst W, Drescher U (2001) A role for the EphA family in the topographic targeting of vomeronasal axons. Development 128:895–906

    CAS  PubMed  Google Scholar 

  • Krol J, Loedige I, Filipowicz W (2010) The widespread regulation of microRNA biogenesis, function and decay. Nat Rev Genet 11:597–610

    CAS  PubMed  Google Scholar 

  • Liu K, Lu Y, Lee JK, et al. (2010) PTEN deletion enhances the regenerative ability of adult corticospinal neurons. Nat Neurosci 13:1075–1081

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Marquardt T, Shirasaki R, Ghosh S, et al. (2005) Coexpressed EphA receptors and ephrin-a ligands mediate opposing actions on growth cone navigation from distinct membrane domains. Cell 121:127–139

    Article  CAS  PubMed  Google Scholar 

  • Martinez A, Soriano E (2005) Functions of ephrin/Eph interactions in the development of the nervous system: emphasis on the hippocampal system. Brain Res Brain Res Rev 49:211–226

    Article  CAS  PubMed  Google Scholar 

  • McLaughlin T, Hindges R, O’Leary DD (2003) Regulation of axial patterning of the retina and its topographic mapping in the brain. Curr Opin Neurobiol 13:57–69

    Article  CAS  PubMed  Google Scholar 

  • Müller J, Ossig C, Greiner JF, et al. (2015) Intrastriatal transplantation of adult human neural crest-derived stem cells improves functional outcome in parkinsonian rats. Stem Cells Transl Med 4:31–43

    Article  PubMed  Google Scholar 

  • Murai KK, Nguyen LN, Irie F, Yamaguchi Y, Pasquale EB (2003) Control of hippocampal dendritic spine morphology through ephrin-A3/EphA4 signaling. Nat Neurosci 6:153–160

    Article  CAS  PubMed  Google Scholar 

  • Natera-Naranjo O, Aschrafi A, Gioio AE, Kaplan BB (2010) Identification and quantitative analyses of microRNAs located in the distal axons of sympathetic neurons. RNA 16:1516–1529

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Pittman A, Chien CB (2002) Understanding dorsoventral topography: backwards and forwards. Neuron 35:409–411

    Article  CAS  PubMed  Google Scholar 

  • Remy P (2014) Biotherapies for Parkinson disease. Rev Neurol (Paris) 170:763–769

  • Rhee YH, Ko JY, Chang MY, et al. (2011) Protein-based human iPS cells efficiently generate functional dopamine neurons and can treat a rat model of Parkinson disease. J Clin Invest 121:2326–2335

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Schratt G (2009) microRNAs at the synapse. Nat Rev Neurosci 10:842–849

    Article  CAS  PubMed  Google Scholar 

  • Seiradake E, Schaupp A, del Toro RD, et al. (2013) Structurally encoded intraclass differences in EphA clusters drive distinct cell responses. Nat Struct Mol Biol 20:958–964

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shin ES, Hwang O, Hwang YS, Suh JK, Chun YI, Jeon SR (2014) Enhanced efficacy of human brain-derived neural stem cells by transplantation of cell aggregates in a rat model of Parkinson’s disease. J Korean Neurosurg Soc 56:383–389

    Article  PubMed  PubMed Central  Google Scholar 

  • Stein E, Savaskan NE, Ninnemann O, Nitsch R, Zhou R, Skutella T (1999) A role for the Eph ligand ephrin-A3 in entorhino-hippocampal axon targeting. J Neurosci 19:8885–8893

    CAS  PubMed  Google Scholar 

  • Vanderhaeghen P, Lu Q, Prakash N, et al. (2000) A mapping label required for normal scale of body representation in the cortex. Nat Neurosci 3:358–365

    Article  CAS  PubMed  Google Scholar 

  • Vo N, Klein ME, Varlamova O, et al. (2005) A cAMP-response element binding protein-induced microRNA regulates neuronal morphogenesis. Proc Natl Acad Sci U S A 102:16426–16431

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wahl S, Barth H, Ciossek T, Aktories K, Mueller BK (2000) Ephrin-A5 induces collapse of growth cones by activating rho and rho kinase. J Cell Biol 149:263–270

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wang HU, Anderson DJ (1997) Eph family transmembrane ligands can mediate repulsive guidance of trunk neural crest migration and motor axon outgrowth. Neuron 18:383–396

    Article  CAS  PubMed  Google Scholar 

  • Wilkinson DG (2001) Multiple roles of EPH receptors and ephrins in neural development. Nat Rev Neurosci 2:155–164

    Article  CAS  PubMed  Google Scholar 

  • Xu B, Li S, Brown A, Gerlai R, Fahnestock M, Racine RJ (2003) EphA/ephrin-a interactions regulate epileptogenesis and activity-dependent axonal sprouting in adult rats. Mol Cell Neurosci 24:984–999

    Article  CAS  PubMed  Google Scholar 

  • Yu JY, Chung KH, Deo M, Thompson RC, Turner DL (2008) MicroRNA miR-124 regulates neurite outgrowth during neuronal differentiation. Exp Cell Res 314:2618–2633

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Yue X, Dreyfus C, Kong TA, Zhou R (2008) A subset of signal transduction pathways is required for hippocampal growth cone collapse induced by ephrin-A5. Dev Neurobiol 68:1269–1286

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Correspondence to Dian-Shuai Gao.

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Ting Wang and Jing Chen contributed equally to this article.

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Wang, T., Chen, J., Tang, CX. et al. Inverse Expression Levels of EphrinA3 and EphrinA5 Contribute to Dopaminergic Differentiation of Human SH-SY5Y Cells. J Mol Neurosci 59, 483–492 (2016). https://doi.org/10.1007/s12031-016-0759-y

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