title: The canonical Wnt pathway regulates the metastasis-promoting mucin MUC4 in pancreatic ductal adenocarcinoma.
abstract: Aberrant Wnt signaling frequently occurs in pancreatic cancer (PC) and contributes to disease progression/metastases. Likewise, the transmembrane-mucin MUC4 is expressed de novo in early pancreatic intraepithelial neoplasia (PanINs) and incrementally increases with PC progression, contributing to metastasis. To determine the mechanism of MUC4 upregulation in PC, we examined factors deregulated in early PC progression, such as Wnt/β-catenin signaling. MUC4 promoter analysis revealed the presence of three putative TCF/LEF-binding sites, leading us to hypothesize that MUC4 can be regulated by β-catenin. Immunohistochemical (IHC) analysis of rapid autopsy PC tissues showed a correlation between MUC4 and cytosolic/nuclear β-catenin expression. Knock down (KD) of β-catenin in CD18/HPAF and T3M4 cell lines resulted in decreased MUC4 transcript and protein. Three MUC4 promoter luciferase constructs, p3778, p3000, and p2700, were generated. The construct p3778, encompassing the entire MUC4 promoter, elicited increased luciferase activity in the presence of stabilized β-catenin. Mutation of the TCF/LEF site closest to the transcription start site (i.e., -2629/-2612) and furthest from the start site (i.e., -3425/-3408) reduced MUC4 promoter luciferase activity. Transfection with dominant negative TCF4 decreased MUC4 transcript and protein levels. Chromatin immunoprecipitation confirmed enrichment of β-catenin on -2629/-2612 and -3425/-3408 of the MUC4 promoter in CD18/HPAF. Functionally, CD18/HPAF and T3M4 β-catenin KD cells showed decreased migration and decreased Vimentin, N-cadherin, and pERK1/2 expression. Tumorigenicity studies in athymic nude mice showed CD18/HPAF β-catenin KD cells significantly reduced primary tumor sizes and metastases compared to scrambled control cells. We show for the first time that β-catenin directly governs MUC4 in PC.
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center (UNMC), Omaha, NE 68198-5870, USA.
authors.forename: Priya
authors.initials: P
authors.lastname: Pai
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center (UNMC), Omaha, NE 68198-5870, USA.
authors.forename: Satyanarayana
authors.initials: S
authors.lastname: Rachagani
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center (UNMC), Omaha, NE 68198-5870, USA.
authors.forename: Imayavaramban
authors.initials: I
authors.lastname: Lakshmanan
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center (UNMC), Omaha, NE 68198-5870, USA.
authors.forename: Muzafar A
authors.initials: MA
authors.lastname: Macha
authors.affiliation: Department of Pathology and Microbiology, UNMC, Omaha, NE 68198-5900, USA.
authors.forename: Yuri
authors.initials: Y
authors.lastname: Sheinin
authors.affiliation: Department of Biostatistics, UNMC College of Public Health, UNMC, Omaha, NE 68198-4375, USA.
authors.forename: Lynette M
authors.initials: LM
authors.lastname: Smith
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center (UNMC), Omaha, NE 68198-5870, USA; Fred and Pamela Buffett Cancer Center, UNMC, Omaha, NE 68198, USA.
authors.forename: Moorthy P
authors.initials: MP
authors.lastname: Ponnusamy
authors.affiliation: Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center (UNMC), Omaha, NE 68198-5870, USA; Eppley Institute for Research in Cancer and Allied Diseases, UNMC, Omaha, NE 68198-5950, USA; Fred and Pamela Buffett Cancer Center, UNMC, Omaha, NE 68198, USA. Electronic address: sbatra@unmc.edu.
authors.forename: Surinder K
authors.initials: SK
authors.lastname: Batra
mesh_terms: D000818:Animals; D015197:Carcinogenicity Tests; D021441:Carcinoma, Pancreatic Ductal / Q000378:metabolism / Q000556:secondary*; D045744:Cell Line, Tumor; D005260:Female; D055785:Gene Knockdown Techniques; D006801:Humans; D051379:Mice; D008819:Mice, Nude; D055264:Mucin-4 / Q000235:genetics / Q000378:metabolism*; D009362:Neoplasm Metastasis; D010190:Pancreatic Neoplasms / Q000378:metabolism / Q000473:pathology*; D011401:Promoter Regions, Genetic / Q000235:genetics; D058118:Transcription Factor 7-Like 2 Protein / Q000378:metabolism*; D015854:Up-Regulation; D060449:Wnt Signaling Pathway; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism*
chemical_list: C495270:CTNNB1 protein, human; C524658:MUC4 protein, human; D055264:Mucin-4; C544894:TCF7L2 protein, human; D058118:Transcription Factor 7-Like 2 Protein; D051176:beta Catenin
keywords: MUC4; Pancreatic cancer; Wnt; β-Catenin