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The role of KCNQ1 in mouse and human gastrointestinal cancers.

Than BL, Goos JA, Sarver AL, O'Sullivan MG, Rod A, Starr TK, Fijneman RJ, Meijer GA, Zhao L, Zhang Y, Largaespada DA, Scott PM, Cormier RT - Oncogene (2013)

Bottom Line: We also found genes implicated in inflammation and in cellular detoxification.Pathway analysis using Ingenuity Pathway Analysis and Gene Set Enrichment Analysis confirmed the importance of these gene clusters and further identified significant overlap with genes regulated by MUC2 and CFTR, two important regulators of intestinal homeostasis.Results showed that low expression of KCNQ1 expression was significantly associated with poor overall survival.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Biomedical Sciences, University of Minnesota Medical School, Duluth, MN, USA [2] Toxicology Graduate Program, University of Minnesota, Duluth, MN, USA.

ABSTRACT
Kcnq1, which encodes for the pore-forming α-subunit of a voltage-gated potassium channel, was identified as a gastrointestinal (GI) tract cancer susceptibility gene in multiple Sleeping Beauty DNA transposon-based forward genetic screens in mice. To confirm that Kcnq1 has a functional role in GI tract cancer, we created Apc(Min) mice that carried a targeted deletion mutation in Kcnq1. Results demonstrated that Kcnq1 is a tumor suppressor gene as Kcnq1 mutant mice developed significantly more intestinal tumors, especially in the proximal small intestine and colon, and some of these tumors progressed to become aggressive adenocarcinomas. Gross tissue abnormalities were also observed in the rectum, pancreas and stomach. Colon organoid formation was significantly increased in organoids created from Kcnq1 mutant mice compared with wild-type littermate controls, suggesting a role for Kcnq1 in the regulation of the intestinal crypt stem cell compartment. To identify gene expression changes due to loss of Kcnq1, we carried out microarray studies in the colon and proximal small intestine. We identified altered genes involved in innate immune responses, goblet and Paneth cell function, ion channels, intestinal stem cells, epidermal growth factor receptor and other growth regulatory signaling pathways. We also found genes implicated in inflammation and in cellular detoxification. Pathway analysis using Ingenuity Pathway Analysis and Gene Set Enrichment Analysis confirmed the importance of these gene clusters and further identified significant overlap with genes regulated by MUC2 and CFTR, two important regulators of intestinal homeostasis. To investigate the role of KCNQ1 in human colorectal cancer (CRC), we measured protein levels of KCNQ1 by immunohistochemistry in tissue microarrays containing samples from CRC patients with liver metastases who had undergone hepatic resection. Results showed that low expression of KCNQ1 expression was significantly associated with poor overall survival.

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qRT-PCR gene expression analysis of mouse proximal small intestineEach gene sample was run in triplicate and gene expression was normalized to the expression of 18S. Data are presented as the mean fold change +/− SD. Each bar represents the mean and standard error of multiple experiments that measured fold differences in the mRNA expression in proximal small intestine tissue isolated from adult (~100 d) littermate and gender matched pairs of Apc+/+Kcnq1−/− and Kcnq1+/+ mice. mRNAs were isolated from 1 cm sections of proximal small intestine from the same region for all mice. At least two matched pairs of mRNAs were tested for each gene with most genes tested in at least three matched pairs of mRNAs. To be included in this figure genes showed a mean fold difference was at least 1.5. In all cases the direction of changes in gene expression confirmed microarray data. * P<0.05.
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Figure 3: qRT-PCR gene expression analysis of mouse proximal small intestineEach gene sample was run in triplicate and gene expression was normalized to the expression of 18S. Data are presented as the mean fold change +/− SD. Each bar represents the mean and standard error of multiple experiments that measured fold differences in the mRNA expression in proximal small intestine tissue isolated from adult (~100 d) littermate and gender matched pairs of Apc+/+Kcnq1−/− and Kcnq1+/+ mice. mRNAs were isolated from 1 cm sections of proximal small intestine from the same region for all mice. At least two matched pairs of mRNAs were tested for each gene with most genes tested in at least three matched pairs of mRNAs. To be included in this figure genes showed a mean fold difference was at least 1.5. In all cases the direction of changes in gene expression confirmed microarray data. * P<0.05.

Mentions: We conducted cDNA microarray expression studies using tissue from distal colon and the proximal quarter of the small intestine of Apc+/+ Kcnq1−/− mice. Overall, more than 400 genes showed a greater than 1.5 fold change in expression, and greater than 200 of these genes also had a P value of < 0.05 (Supp. Tables 1&2). Expression of 20 genes was confirmed by RT-PCR (Figures 2 & 3). Target genes were clustered into several functional groups: innate immune response/goblet & Paneth cell function (Ang4, Retnlb, Clps, Pnliprp2, Clca3, Muc2, Reg3a, Reg3b, Reg3g, Car1), ion channels (Trpv6, Slc12a8, Slc30a10, Aqp7, Aqp1, Aqp8, Aqp4, Clca3, Clca6), mucins (Muc2, Muc13, Muc4, Muc3), growth regulatory signaling pathways (Areg, Egr1, Fos, Ccdn1), cancer cell migration (Mmp9, Mmp15, S100a14, Mt1), apoptosis (Casp14, Casp2, Bcl6), inflammation, mediated by mechanisms such as detoxification & stress responses (Cyp2c55, Gstk1, Sirt1, Sirt3, Aldh1a1, Aldh1b1, Aldh2, Mt1, Gstm2), the adaptive immune response, and intestinal stem cell-related genes (Clca4, Aqp4, Aldh1a1, Olfm4), consistent with a role for Kcnq1 in the intestinal cell compartment22. Interestingly, while loss of Kcnq1 showed a strong effect on colon secretory cell genes, there was no change in colon tissues in the secretory cell populations or cell proliferation (Supp. Fig. 4).


The role of KCNQ1 in mouse and human gastrointestinal cancers.

Than BL, Goos JA, Sarver AL, O'Sullivan MG, Rod A, Starr TK, Fijneman RJ, Meijer GA, Zhao L, Zhang Y, Largaespada DA, Scott PM, Cormier RT - Oncogene (2013)

qRT-PCR gene expression analysis of mouse proximal small intestineEach gene sample was run in triplicate and gene expression was normalized to the expression of 18S. Data are presented as the mean fold change +/− SD. Each bar represents the mean and standard error of multiple experiments that measured fold differences in the mRNA expression in proximal small intestine tissue isolated from adult (~100 d) littermate and gender matched pairs of Apc+/+Kcnq1−/− and Kcnq1+/+ mice. mRNAs were isolated from 1 cm sections of proximal small intestine from the same region for all mice. At least two matched pairs of mRNAs were tested for each gene with most genes tested in at least three matched pairs of mRNAs. To be included in this figure genes showed a mean fold difference was at least 1.5. In all cases the direction of changes in gene expression confirmed microarray data. * P<0.05.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3935979&req=5

Figure 3: qRT-PCR gene expression analysis of mouse proximal small intestineEach gene sample was run in triplicate and gene expression was normalized to the expression of 18S. Data are presented as the mean fold change +/− SD. Each bar represents the mean and standard error of multiple experiments that measured fold differences in the mRNA expression in proximal small intestine tissue isolated from adult (~100 d) littermate and gender matched pairs of Apc+/+Kcnq1−/− and Kcnq1+/+ mice. mRNAs were isolated from 1 cm sections of proximal small intestine from the same region for all mice. At least two matched pairs of mRNAs were tested for each gene with most genes tested in at least three matched pairs of mRNAs. To be included in this figure genes showed a mean fold difference was at least 1.5. In all cases the direction of changes in gene expression confirmed microarray data. * P<0.05.
Mentions: We conducted cDNA microarray expression studies using tissue from distal colon and the proximal quarter of the small intestine of Apc+/+ Kcnq1−/− mice. Overall, more than 400 genes showed a greater than 1.5 fold change in expression, and greater than 200 of these genes also had a P value of < 0.05 (Supp. Tables 1&2). Expression of 20 genes was confirmed by RT-PCR (Figures 2 & 3). Target genes were clustered into several functional groups: innate immune response/goblet & Paneth cell function (Ang4, Retnlb, Clps, Pnliprp2, Clca3, Muc2, Reg3a, Reg3b, Reg3g, Car1), ion channels (Trpv6, Slc12a8, Slc30a10, Aqp7, Aqp1, Aqp8, Aqp4, Clca3, Clca6), mucins (Muc2, Muc13, Muc4, Muc3), growth regulatory signaling pathways (Areg, Egr1, Fos, Ccdn1), cancer cell migration (Mmp9, Mmp15, S100a14, Mt1), apoptosis (Casp14, Casp2, Bcl6), inflammation, mediated by mechanisms such as detoxification & stress responses (Cyp2c55, Gstk1, Sirt1, Sirt3, Aldh1a1, Aldh1b1, Aldh2, Mt1, Gstm2), the adaptive immune response, and intestinal stem cell-related genes (Clca4, Aqp4, Aldh1a1, Olfm4), consistent with a role for Kcnq1 in the intestinal cell compartment22. Interestingly, while loss of Kcnq1 showed a strong effect on colon secretory cell genes, there was no change in colon tissues in the secretory cell populations or cell proliferation (Supp. Fig. 4).

Bottom Line: We also found genes implicated in inflammation and in cellular detoxification.Pathway analysis using Ingenuity Pathway Analysis and Gene Set Enrichment Analysis confirmed the importance of these gene clusters and further identified significant overlap with genes regulated by MUC2 and CFTR, two important regulators of intestinal homeostasis.Results showed that low expression of KCNQ1 expression was significantly associated with poor overall survival.

View Article: PubMed Central - PubMed

Affiliation: 1] Department of Biomedical Sciences, University of Minnesota Medical School, Duluth, MN, USA [2] Toxicology Graduate Program, University of Minnesota, Duluth, MN, USA.

ABSTRACT
Kcnq1, which encodes for the pore-forming α-subunit of a voltage-gated potassium channel, was identified as a gastrointestinal (GI) tract cancer susceptibility gene in multiple Sleeping Beauty DNA transposon-based forward genetic screens in mice. To confirm that Kcnq1 has a functional role in GI tract cancer, we created Apc(Min) mice that carried a targeted deletion mutation in Kcnq1. Results demonstrated that Kcnq1 is a tumor suppressor gene as Kcnq1 mutant mice developed significantly more intestinal tumors, especially in the proximal small intestine and colon, and some of these tumors progressed to become aggressive adenocarcinomas. Gross tissue abnormalities were also observed in the rectum, pancreas and stomach. Colon organoid formation was significantly increased in organoids created from Kcnq1 mutant mice compared with wild-type littermate controls, suggesting a role for Kcnq1 in the regulation of the intestinal crypt stem cell compartment. To identify gene expression changes due to loss of Kcnq1, we carried out microarray studies in the colon and proximal small intestine. We identified altered genes involved in innate immune responses, goblet and Paneth cell function, ion channels, intestinal stem cells, epidermal growth factor receptor and other growth regulatory signaling pathways. We also found genes implicated in inflammation and in cellular detoxification. Pathway analysis using Ingenuity Pathway Analysis and Gene Set Enrichment Analysis confirmed the importance of these gene clusters and further identified significant overlap with genes regulated by MUC2 and CFTR, two important regulators of intestinal homeostasis. To investigate the role of KCNQ1 in human colorectal cancer (CRC), we measured protein levels of KCNQ1 by immunohistochemistry in tissue microarrays containing samples from CRC patients with liver metastases who had undergone hepatic resection. Results showed that low expression of KCNQ1 expression was significantly associated with poor overall survival.

Show MeSH
Related in: MedlinePlus