The International Journal of Biochemistry & Cell Biology 61 (2015) 53–62 Contents lists available at ScienceDirect The International Journal of Biochemistry & Cell Biology jo ur nal home page: www.elsevier.com/locate/biocel PTPN21 exerts pro-neuronal survival and neuritic elongation via ErbB4/NRG3 signaling Janice Hiu-Chor Plani-Lam a , Tai-Cheong Chow a , Kam-Leung Siu a , Wing Hin Chau a , Ming-Him James Ng a,b , Suying Bao a , Cheung Toa Ng a , Pak Sham c,d , Daisy Kwok-Yan Shum a , Evan Ingley e , Dong-Yan Jin a , You-Qiang Song a,* a Department of Biochemistry, University of Hong Kong, 21 Sassoon Road, Hong Kong, China b Poison Treatment Centre, Department of Medicine and Therapeutics, Prince of Wales Hospital, Shatin, Hong Kong, China c Department of Psychiatry, University of Hong Kong, 21 Sassoon Road, Hong Kong, China d Centre for Genomic Sciences, University of Hong Kong, 5 Sassoon Road, Hong Kong, China e Harry Perkins Institute of Medical Research, QEII Medical Centre, Nedlands and Centre for Medical Research, The University of Western Australia, Crawley, Western Australia, 6009, Australia a r t i c l e i n f o Article history: Received 11 October 2014 Received in revised form 27 January 2015 Accepted 3 February 2015 Available online 11 February 2015 Keywords: PTPN21 NRG3 ErbB4 Pro-neuronal survival Neuritic elongation a b s t r a c t Although expression quantitative trait locus, eQTL, serves as an explicit indicator of gene–gene asso- ciations, challenges remain to disentangle the mechanisms by which genetic variations alter gene expression. Here we combined eQTL and molecular analyses to identify an association between two seemingly non-associated genes in brain expression data from BXD inbred mice, namely Ptpn21 and Nrg3. Using biotinylated receptor tracking and immunoprecipitation analyses, we determined that PTPN21 de-phosphorylates the upstream receptor tyrosine kinase ErbB4 leading to the up-regulation of its down- stream signaling. Conversely, kinase-dead ErbB4 (K751R) or phosphatase-dead PTPN21 (C1108S) mutants impede PTPN21-dependent signaling. Furthermore, PTPN21 also induced Elk-1 activation in embryonic cortical neurons and a novel Elk-1 binding motif was identified in a region located 1919 bp upstream of the NRG3 initiation codon. This enables PTPN21 to promote NRG3 expression through Elk-1, which pro- vides a biochemical mechanism for the PTPN21–NRG3 association identified by eQTL. Biologically, PTPN21 positively influences cortical neuronal survival and, similar to Elk-1, it also enhances neuritic length. Our combined approaches show for the first time, a link between NRG3 and PTPN21 within a signaling cas- cade. This may explain why these two seemingly unrelated genes have previously been identified as risk genes for schizophrenia. © 2015 Elsevier Ltd. All rights reserved. 1. Introduction The neurotrophic factor Neuregulin 3 (NRG3), has been iden- tified as a risk gene for Schizophrenia in multiple genome-wide association studies in Ashkenazi Jewish (Chen et al., 2009), Abbreviations: NRG3, Neuregulin 3; eQTL, expression quantitative trait locus; SNPs, single nucleotide polymorphisms; PTPN21, protein tyrosine phosphatase non- receptor 21; GWAS, genome-wide association study; RTK, receptor tyrosine kinase; FAK, focal adhesion kinase; KIF1C, tyrosine-phosphorylated kinesin-like Protein; Etk, Tec tyrosine kinase; Elk-1, ETS domain-containing protein; ErbB4, V-erb-b2 avian erythroblastic leukemia viral oncogene homolog 4; IP, immunoprecipitation; p-Tyr, tyrosine phosphatase; RT-qPCR, quantitative reverse transcriptase-PCR. * Corresponding author at: Room L3-63, Department of Biochemistry, University of Hong Kong, 21 Sassoon Road, Hong Kong, China. Tel.: +852 2819 9245; fax: +852 2855 1254. E-mail address: songy@hku.hk (Y.-Q. Song). Australian (Morar et al., 2011), German (Meier et al., 2013) and Han Chinese (Wang et al., 2008) populations. The presence of other important genetic components is indicated by numerous other genomic studies, including a recent GWAS meta-analyses that identified protein tyrosine phosphatase non-receptor 21 (PTPN21) as a potential risk gene for schizophrenia (Chen et al., 2011). This genetic study identified two missense non-synonymous sin- gle nucleotide polymorphisms (SNPs) in PTPN21, rs2401751 and rs2274736, which change the size and alter the hydrophobicity of residues that may impact enzyme activity. Intriguingly, an indepen- dent quantitative trait locus (eQTL), of brain in BXD recombinant inbred mice also identified that the genetic variants of PTPN21 mediate Nrg3 expression in mouse neurons. This eQTLs mapping method revealed association between genetic polymorphisms vari- ation and mRNA expression variation (Michaelson et al., 2009). Since the results of both eQTL and multiple schizophrenia genome- wide association study (GWAS) studies are significantly in support http://dx.doi.org/10.1016/j.biocel.2015.02.003 1357-2725/© 2015 Elsevier Ltd. All rights reserved.