SPLUNC1 Regulates Cell Progression and Apoptosis through the miR-141-PTEN/p27 Pathway, but Is Hindered by LMP1

but Is Hindered by LMP1. PLoS ONE 8(3): e56929. doi:10.1371/journal.pone.0056929 SPLUNC1 Regulates Cell Progression and Apoptosis through the miR-141-PTEN/p27 Pathway, but Is Hindered by LMP1 Pan Chen Xiaofang Guo Houde Zhou Wenling Zhang Zhaoyang Zeng Qianjin Liao Xiayu Li Bo Xiang Jianbo Yang Jian Ma Ming Zhou Shuping Peng Juanjuan Xiang Xiaoling Li Colvin Wanshura LE Wei Xiong James B. McCarthy Guiyuan Li Rajeev Samant, University of Alabama at Birmingham, United States of America Little is known about the role of the host defensive protein short palate, lung and nasal epithelium clone 1 (SPLUNC1) in the carcinogenesis of nasopharyngeal carcinoma (NPC). Here we report that SPLUNC1 plays a role at a very early stage of NPC carcinogenesis. SPLUNC1 regulates NPC cell proliferation, differentiation and apoptosis through miR-141, which in turn regulates PTEN and p27 expression. This signaling axis is negatively regulated by the EBV-coded gene LMP1. Therefore we propose that SPLUNC1 suppresses NPC tumor formation and its inhibition by LMP1 provides a route for NPC tumorigenesis. - Funding: This study was supported by grants from the National High Technology Research and Development Program of China (2012AA02A206), the National Natural Science Foundation of China (91229122, 81071644, 81101509, 81171934, 81172189, 81171930, 81272298, 81272254 and 30971147), the 111 Project (1112-12) and the Hunan Province Natural Sciences Foundation of China (10JJ7003). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. . These authors contributed equally to this work. Nasopharyngeal carcinoma (NPC) is a nonlymphomatous, squamous cell carcinoma that occurs in the epithelial lining of the nasopharynx [1,2]. NPC represents a significant clinical challenge in southern China and Southeast Asia, with an incidence of 1550 per 100,000 [3]. NPC development is a multistage process influenced by complex interactions between genetic and environmental factors [4]. Chronic infection with EpsteinBarr virus (EBV) or other pathogens has been considered one of the most important factors NPC development [5,6]. The EBVencoded latent membrane protein 1 (LMP1) is an oncogene that is critically involved in the EBV-driven immortalization of B cells in vitro, and is able to transform rodent fibroblast and human cell lines in culture [7]. More than 90% of the adult population of the world have been infected by EBV, and although they become lifelong carriers of the virus, only about 0.03% of them develop NPC [8], which indicates that other factors exist to protect the EBV-infected nasopharyngeal epithelium against NPC. Innate immune responses in epithelial cells provide a vital source of host defense molecules to protect against nasopharyngeal epithelial infection. These host defense molecules are natural immunobarriers against environmental threats with various mechanisms to bind, transport, cleave, or degrade microbes and their endotoxic byproducts [9], which decrease the risk of epithelial damage and the development of cancer. Short palate, lung and nasal epithelium clone 1 (SPLUNC1) was originally found to be downregulated in NPC and exhibits host defense properties [10,11]. SPLUNC1 is secreted by large airway epithelial cells and has been shown to possess antimicrobial and antiinflammatory functions [12,13]. SPLUNC1 is able to induce apoptosis of EBV-infected B lymphocytes, and polymorphisms in the SPLUNC1 locus are associated with susceptibility to NPC [14]. SPLUNC1 also serves to protect the sodium channel protein (ENaC) from proteolytic cleavage, thus regulating airway surface liquid volume [15]. Effective maintenance of mucosal liquid volume, mediated by appropriate expression of SPLUNC1, is thought to promote mucociliary clearance of microorganisms from the airway [16]. While the involvement of SPLUNC1 in host defensive protein is well-delineated, its involvement in the tumorigenesis of NPC remains unclear. Previous differential analysis of microRNA (miRNA) expression profiles after re-expression of SPLUNC1 in NPC cells showed that SPLUNC1 could decrease miR-141 expression in the highly tumorigenic and metastatic 58F NPC cells. PTEN has been validated as a miR-141 target by 39-untranslated region (39-UTR) luciferase reporter assays [17]. While the extent of PTEN involvement in NPC tumorigenesis is an area of current investigation, loss of PTEN function through deletion, mutation, and/or decreased expression has been found in numerous human sporadic cancers [18,19] and in hereditary cancer syndromes [20,21]. Furthermore, activation of PI3K/AKT and other signalling pathways by LMP1 leads to enhance NPC cell growth and migration [2225]. Together, these data indicate that crosstalk between the SPLUNC1-miR-141-PTEN and LMP1Akt signaling axes may be important in NPC development and progre (...truncated)