Clostridium perfringens enterotoxin induces claudin-4 to activate YAP in oral squamous cell carcinomas

// Chie Nakashima 1 , 2 , Kazuhiko Yamamoto 2 , Shingo Kishi 1 , Takamitsu Sasaki 1 , Hitoshi Ohmori 1 , Rina Fujiwara-Tani 1 , Shiori Mori 1 , Isao Kawahara 1 , Yukiko Nishiguchi 1 , Takuya Mori 1 , Masuo Kondoh 3 , Yi Luo 4 , Tadaaki Kirita 2 and Hiroki Kuniyasu 1 1 Department of Molecular Pathology, Nara Medical University, Kashihara, Nara 634-8521, Japan 2 Department of Oral and Maxillofacial Surgery, Nara Medical University, Kashihara, Nara 634-8522, Japan 3 Drug Innovation Center, Graduate School of Pharmaceutical Sciences, Osaka University, Suita, Osaka 565-0871, Japan 4 Key Laboratory of Neuroregeneration of Jiangsu and Ministry of Education, Co-Innovation Center of Neuroregeneration, Nantong University, Nantong, Jiangsu Province 226001, China Correspondence to: Yi Luo, email: lynantong@hotmail.com Tadaaki Kirita, email: tkirita@naramed-u.ac.jp Hiroki Kuniyasu, email: cooninh@zb4.so-net.ne.jp Keywords: clostridium perfringens enterotoxin; claudin-4; YAP; Hippo signal; oral squamous cell carcinoma; Pathology Received: November 11, 2019     Accepted: December 21, 2019     Published: January 28, 2020 ABSTRACT Claudin (CLDN)-4 expression has been associated with malignancy in various cancers. When CLDN4 expression was examined in oral squamous cell carcinoma (OSCC), 22 out of 57 (39%) cases showed immunoreactivity in the nucleus. Nuclear CLDN4-positive cases showed a stronger correlation with cancer progression than the negative cases. Intratumoral anaerobic bacterial DNA examination revealed nuclear CLDN4 expression in 81% of Clostridium perfringens -positive cases. Treatment of human oral squamous cell carcinoma cell lines HSC3 and HSC4 with Clostridium perfringens enterotoxin (CPE), induced CLDN4 nuclear translocation to enhance epithelial-mesenchymal transition (EMT), stemness, cell proliferation and invasive ability. In addition, CPE treatment suppressed phosphorylation of yes-associated protein-1 (YAP1) and promoted YAP1 nuclear translocation, resulting in increased expression of YAP1 target genes; cyclin D1 and connective tissue growth factor. Moreover, it was revealed that the complex of YAP1, CLDN4 and zona occludens-2 (ZO-2) was formed by CPE treatment, further suppressing YAP1 phosphorylation by LATS1 and activating it. Thus YAP activation in OSCC was regarded important in promoting malignant phenotypes. Our research suggested that the control of oral anaerobic bacteria may suppress YAP activation and in turn tumor progression.