Stable expression of four different ErbB4 isoforms - differing in their juxtamembrane region (JMa/JMb) and C terminus (cyt1/cyt2) - confers to neural progenitor cells (ST14A) the ability to migrate following neuregulin 1 (NRG1) stimulation, although with different intensity: JMa-cyt2 and JMb-cyt1 expression providing the higher migratory activity. To identify genes regulated by ErbB4 and involved in neuronal migration, we performed microarray analysis to compare RNA obtained from mock cells with RNA obtained from ErbB4 JMa-cyt2 expressing cells, both stimulated with NRG1. The identified regulated genes belong to many different categories; nevertheless, we focused our attention on genes involved in migration, and in particular in cytoskeleton organization. By quantitative real time RT-PCR we analyzed the expression level of these genes in stable cell clones, each expressing one of the four different ErbB4 isoforms, in the presence or not of NRG1 stimulation. Our results show that some candidate genes display an expression pattern which is not correlated with ErbB4 mediated migration activity, whereas other genes seem to be influenced by NRG1 stimulation of ErbB4, being down- or up-regulated. Only a few genes display an expression pattern which is correlated with ErbB4 mediated migration activity. We focused our attention on Eps8 (EGF receptor pathway substrate 8), which is a gene involved in the regulation of actin dynamics. Here we show, that stable cell clones endowed with the higher migratory activity, display a higher expression of Eps8 mRNA and protein. Two different Eps8 isoforms have been described (97- and 68-kDa) differing for their N terminus domain. By transient expression of different siRNAs, we silenced both Eps8 isoforms or only the full length, and we demonstrated, by Transwell assays, that Eps8 97kDa isoform is necessary to mediate NRG1 induced migration. Financial support from Regione Piemonte and MUR.
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