Overexpression of p54 nrb /NONO induces differential EPHA6 splicing and contributes to castration-resistant prostate cancer growth

// Ryuji Yamamoto 1 , Tsuyoshi Osawa 1 , Yusuke Sasaki 1 , Shogo Yamamoto 2 , Motonobu Anai 1 , Kouji Izumi 3 , Yoshihiro Matsumura 4 , Juro Sakai 4, 5 , Hiroyuki Aburatani 2 , Atsushi Mizokami 3 , Tatsuhiko Kodama 1 and Toshiya Tanaka 1 1 Laboratory for Systems Biology and Medicine (LSBM), Research Center for Advanced Science and Technology (RCAST), The University of Tokyo, Tokyo 153-8904, Japan 2 Division of Genome Science, Research Center for Advanced Science and Technology (RCAST), The University of Tokyo, Tokyo 153-8904, Japan 3 Department of Integrative Cancer Therapy and Urology, Division of Cancer Medicine, Graduate School of Medical Science, Kanazawa University, Kanazawa, Ishikawa 920-8641, Japan 4 Division of Metabolic Medicine, Research Center for Advanced Science and Technology (RCAST), The University of Tokyo, Tokyo 153-8904, Japan 5 Division of Molecular Physiology and Metabolism, Tohoku University Graduate School of Medicine, Sendai 980-8574, Japan Correspondence to: Toshiya Tanaka, email: tanaka@lsbm.org Keywords: CRPC; neuroendocrine prostate cancer; p54 nrb /NONO; RNA splicing; EPHA6 Abbreviations: PCa-prostate cancer, CRPC-castration-resistant prostate cancer, LNCaP-SF-LNCaP steroid free Received: August 24, 2017      Accepted: January 02, 2018      Published: January 08, 2018 ABSTRACT The non-POU domain-containing octamer binding protein p54 nrb /NONO is a multifunctional nuclear protein involved in RNA splicing, processing, and transcriptional regulation of nuclear hormone receptors. Through chromosome copy number analysis via whole-exome sequencing, we revealed amplification of the chromosome Xq11.22-q21.33 locus containing the androgen receptor ( AR ) and NONO genes in androgen-independent, castration-resistant prostate cancer (CRPC)-like LNCaP-SF cells. Moreover, NONO was frequently amplified and overexpressed in patients with CRPC. RNA sequencing data revealed that a truncated ephrin type-A receptor 6 ( EPHA6 ) splice variant ( EPHA6-001 ) was overexpressed in LNCaP-SF cells, and knockdown of NONO or EPHA6-001 prevented EPHA6-001 expression and reduced proliferation and invasion by LNCaP-SF cells grown under androgen deprivation conditions. Growth inhibition and differential splicing of EPHA6 mRNA by p54 nrb /NONO were confirmed in gene silencing experiments in 22Rv1 PCa cells. Importantly, NONO knockdown in LNCaP-SF cells led to reduced tumor growth in castrated mice. These findings indicate that p54 nrb /NONO is amplified and overexpressed in CRPC cells and clinical samples, and facilitates CRPC growth by mediating aberrant EPHA6 splicing. We therefore propose that p54 nrb /NONO constitutes a novel and attractive therapeutic target for CRPC.