Oncogene (2012) 31, 3679–3695 & 2012 Macmillan Publishers Limited All rights reserved 0950-9232/12 www.nature.com/onc ORIGINAL ARTICLE Suppression of Tumorigenicity-14, encoding matriptase, is a critical suppressor of colitis and colitis-associated colon carcinogenesis P Kosa, R Szabo, AA Molinolo and TH Bugge Oral and Pharyngeal Cancer Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD, USA Colitis-associated colorectal cancers are an etiologically (collectively, inflammatory bowel disease) with an distinct subgroup of colon cancers that occur in individuals incidence that is proportional to the duration of the suffering from inflammatory bowel disease and arise as a disease. The neoplastic progression of disease-stricken consequence of persistent exposure of hyperproliferative colonic epithelium is believed to be driven by the chronic epithelial stem cells to an inflammatory microenvironment. inflammatory microenvironment, which promotes the An intrinsic defect in the intestinal epithelial barrier has progressive genomic instability of colonic epithelial stem been proposed to be one of several factors that contribute cells by inducing sustained hyperproliferation (regen- to the inappropriate immune response to the commensal erative atypia) and by the continuous presence of high microbiota that underlies inflammatory bowel disease. local concentrations of DNA-damaging agents, such as Matriptase is a membrane-anchored serine protease reactive oxygen species (reviewed in the studies by encoded by Suppression of Tumorigenicity-14 (ST14) that Danese and Mantovani (2010) and Saleh and Trinchieri strengthens the intestinal epithelial barrier by promoting (2011)). tight junction formation. Here, we show that intestinal Although there is considerable debate about the epithelial-specific ablation of St14 in mice causes forma- relative importance of the specific factors that contribute tion of colon adenocarcinoma with very early onset and to the development of inflammatory bowel disease, there high penetrance. Neoplastic progression is preceded by a is a consensus that the disease represents an inappropri- chronic inflammation of the colon that resembles human ate immune response to the commensal microbiota in inflammatory bowel disease and is promoted by the genetically predisposed individuals (reviewed in the commensal microbiota. This study demonstrates that studies by Schreiber et al., 2005; Xavier and Podolsky, inflammation-associated colon carcinogenesis can be 2007; Van Limbergen et al., 2009; Kaser et al., 2010; initiated and promoted solely by an intrinsic intestinal Saleh and Trinchieri, 2011). In this regard, the permeability barrier perturbation, establishes St14 as a contribution of aberrant inflammatory circuits to the critical tumor-suppressor gene in the mouse gastrointest- development of inflammatory bowel disease has been inal tract and adds matriptase to the expanding list clearly established by genetic analysis, including gen- of pericellular proteases with tumor-suppressive functions. ome-wide association studies, which have linked loss-of- Oncogene (2012) 31, 3679–3695; doi:10.1038/onc.2011.545; function mutations or polymorphisms in genes encoding published online 5 December 2011 interleukins, interleukin receptors, chemokine receptors, nucleotide-binding oligomerization domain-like recep- Keywords: inflammatory bowel disease; intestinal tors, Toll-like receptor 4, intelectins and prostaglandin barrier; colon carcinogenesis receptors to ulcerative colitis, to Crohn’s disease or to both. Further support for a principal role of derailed inflammatory circuits is gained from the spontaneous inflammatory bowel disease observed in mice deficient in Introduction various immune effectors (reviewed in the studies by Schreiber et al., 2005; Xavier and Podolsk, 2007; Van Colitis-associated colorectal cancers are etiologically Limbergen et al., 2009; Kaser et al., 2010; Saleh and and molecularly distinct from familial adenomatous Trinchieri, 2011). Much less explored is the importance polyposis coli-associated colorectal cancer, hereditary of individual components of the intestinal epithelial non-polyposis coli colorectal cancer and sporadic color- barrier in preventing inflammatory bowel disease, and ectal cancer. The malignancy occurs in individuals the potential contribution of intrinsic intestinal epithe- suffering from ulcerative colitis or Crohn’s disease lial barrier defects to the development of the syndrome. The clearest indication of the potential importance Correspondence: Dr TH Bugge, Proteases and Tissue Remodeling of primary barrier integrity comes from studies of Section, Oral and Pharyngeal Cancer Branch, National Institute of mice with germline ablation of Muc2 encoding the Dental and Craniofacial Research, National Institutes of Health, 30 major mucin that shields the intestinal epithelium Convent Drive, Room 211, Bethesda, MD 20892, USA. E-mail: [email protected] from direct contact with the microbiota. These Received 6 July 2011; revised 13 October 2011; accepted 27 October mice develop colitis, which may progress to colon 2011; published online 5 December 2011 adenocarcinomas in older animals (Velcich et al., Epithelial barrier defect-induced carcinogenesis P Kosa et al 3680 2002). Additional evidence has been obtained from 2 *** transgenic mice with intestinal epithelial-specific over- expression of myosin light-chain kinase, which display decreased barrier function and increased immune activation, although an inflammatory bowel-like syn- drome did not emerge in the absence of additional immunological challenges (Su et al., 2009). Matriptase is a member of the recently established 1 family of type II transmembrane serine proteases that is *** * * fold change ** ** encoded by the Suppression of Tumorigenicity-14 (ST14) *** *** gene (Kim et al., 1999; Lin et al., 1999; Takeuchi et al., 1999; Tanimoto et al., 2001; Bugge et al., 2009). ST14 was originally proposed to be a colon cancer tumor- suppressor gene, due to its specific downregulation in 0 adenocarcinomas of the colon (Zhang et al., 1998). AIB C D E F G H J K L M N Matriptase is expressed in multiple epithelia of the Figure 1 Matriptase expression is downregulated in human colon integumental, gastrointestinal and urogenital systems, adenomas and adenocarcinomas. Expression of ST14, encoding where it has pleiotropic functions in the differentiation matriptase, in 14 gene expression array studies of human colon or homeostasis of both simple and stratified epithelia, at adenomas and adenocarcinomas. Data are expressed as fold change relative to the corresponding normal tissue. *Po0.05, least in part through the proteolytic activation of the **Po0.01, ***Po0.001. See Supplementary Table 1 for details epithelial sodium channel activator, prostasin/PRSS8 and references. (Szabo and Bugge, 2011). In the simple epithelium of the gastrointestinal tract, a principal function of matriptase is to promote the formation of a paracellular perme- upregulated (Figure 1 and Supplementary Table 1). Of ability barrier, possibly through the posttranslational the 14 studies, study A, which compared gene expression regulation of the composition of claudins that are in colorectal dysplastic adenomatous polyps with the incorporated into the epithelial tight junction complex normal colonic epithelium, was conducted using laser of differentiating intestinal epithelial cells (List et al., capture microdissected tissue (ST14 downregulation, 2009; Buzza et al., 2010). Po0.0006) (Gaspar et al., 2008), and therefore provided Through lineage-specific loss-of-function analysis in the most reliable estimate of ST14 expression in normal mice, we now have examined the role of St14 as a and dysplastic colonic epithelium. tumor-suppressor gene. Interestingly, we found that the selective ablation of St14 from the intestinal epithelium results in the formation of adenocarcinoma of the colon The St14-ablated colonic epithelium undergoes rapid with very early onset and high penetrance. Neoplastic and spontaneous malignant transformation progression occurs in the absence of exposure of animals To specifically explore the functional consequences of to carcinogens or tumor-promoting agents, is preceded intestinal loss of St14 on colon carcinogenesis, we by chronic colonic inflammation that resembles human interbred mice carrying an St14LoxP allele (List et al., inflammatory bowel disease and can be suppressed by 2009) with mice carrying an St14 null allele (St14À) and aggressive antibiotics treatment. This study demon- mice carrying a Cre transgene under the control of the strates that inflammation-associated colon carcinogen- intestinal-specific villin promoter (villin-Cre þ ) (Madison esis can be initiated solely by intrinsic paracellular et al., 2002). This resulted in the generation of villin- permeability barrier perturbations, and establishes that Cre þ /0;St14LoxP/À mice (hereafter termed ‘St14À’ mice) St14 is a critical tumor-suppressor gene in the mouse and their associated littermates villin-Cre þ /0;St14LoxP/ þ , gastrointestinal tract. villin-Cre0;St14LoxP/À and villin-Cre0;St14LoxP/ þ (hereafter termed ‘St14 þ ’ mice). As reported recently (List et al., 2009), this strategy resulted in the efficient deletion of Results matriptase from the entire intestinal tract, as shown by the loss of matriptase immunoreactivity in the colon Meta-analysis of transcriptomes shows decreased (compare Supplementary Figures 1a and b) and the expression of ST14