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MUC1 regulates cyclin D1 gene expression through p120 catenin and β-catenin.

Liu X, Caffrey TC, Steele MM, Mohr A, Singh PK, Radhakrishnan P, Kelly DL, Wen Y, Hollingsworth MA - Oncogenesis (2014)

Bottom Line: We observed a significant enhancement in the activation of cyclin D1 promoter-reporter activity in poorly differentiated Panc1.MUC1F cells that overexpress recombinant MUC1 relative to Panc-1.NEO cells, which express very low levels of endogenous MUC1.Further, MUC1 affected subcellular localization of p120 catenin in association with one of the main effectors of P120 catenin, the transcriptional repressor Kaiso, supporting the hypothesis that p120 catenin relieved transcriptional repression by Kaiso.Our data show MUC1 enhances the activities of both β-catenin and p120 catenin.

View Article: PubMed Central - PubMed

Affiliation: Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha, NE, USA.

ABSTRACT
MUC1 interacts with β-catenin and p120 catenin to modulate WNT signaling. We investigated the effect of overexpressing MUC1 on the regulation of cyclin D1, a downstream target for the WNT/β-catenin signaling pathway, in two human pancreatic cancer cell lines, Panc-1 and S2-013. We observed a significant enhancement in the activation of cyclin D1 promoter-reporter activity in poorly differentiated Panc1.MUC1F cells that overexpress recombinant MUC1 relative to Panc-1.NEO cells, which express very low levels of endogenous MUC1. In stark contrast, cyclin D1 promoter activity was not affected in moderately differentiated S2-013.MUC1F cells that overexpressed recombinant MUC1 relative to S2-013.NEO cells that expressed low levels of endogenous MUC1. The S2-013 cell line was recently shown to be deficient in p120 catenin. MUC1 is known to interact with P120 catenin. We show here that re-expression of different isoforms of p120 catenin restored cyclin D1 promoter activity. Further, MUC1 affected subcellular localization of p120 catenin in association with one of the main effectors of P120 catenin, the transcriptional repressor Kaiso, supporting the hypothesis that p120 catenin relieved transcriptional repression by Kaiso. Thus, full activation of cyclin D1 promoter activity requires β-catenin activation of TCF-lef and stabilization of specific p120 catenin isoforms to relieve the repression of KAISO. Our data show MUC1 enhances the activities of both β-catenin and p120 catenin.

No MeSH data available.


Related in: MedlinePlus

P120 catenin was not detected in S2-013 cells and re-expression of different p120 catenin isoforms in this cells showed distinct subcellular localization. (a) Membrane/cytoplasmic extracts and nuclear extracts from S2-013.NEO, S2-013.MUC1F, Panc-1.NEO, Panc-1.MUC1F were subjected to 10% SDS–PAGE and analyzed by immunoblot (IB) with an anti-p120 catenin mAb. The same blot was striped and reprobed with anti-MUC1.CT antibody. (b) Re-expression of different p120 catenin isoforms were confirmed by western blot using the same p120 catenin antibody and MUC1 antibody. (c and d) Immunofluorescence analysis of re-expression of p120 catenin in S2-013 cells. The green color indicates stains for p120 catenin. Blue indicates 4′-6-diamidino-2-phenylindole (DAPI) staining for nuclei. The arrowhead indicates subcellular localization of p120 catenin in the nucleus.
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fig3: P120 catenin was not detected in S2-013 cells and re-expression of different p120 catenin isoforms in this cells showed distinct subcellular localization. (a) Membrane/cytoplasmic extracts and nuclear extracts from S2-013.NEO, S2-013.MUC1F, Panc-1.NEO, Panc-1.MUC1F were subjected to 10% SDS–PAGE and analyzed by immunoblot (IB) with an anti-p120 catenin mAb. The same blot was striped and reprobed with anti-MUC1.CT antibody. (b) Re-expression of different p120 catenin isoforms were confirmed by western blot using the same p120 catenin antibody and MUC1 antibody. (c and d) Immunofluorescence analysis of re-expression of p120 catenin in S2-013 cells. The green color indicates stains for p120 catenin. Blue indicates 4′-6-diamidino-2-phenylindole (DAPI) staining for nuclei. The arrowhead indicates subcellular localization of p120 catenin in the nucleus.

Mentions: We recently reported that the S2-013 cell line is deficient in expression of p120 catenin, and that re-expression of p120 catenin in these cells stabilized and enhanced steady-state levels of β-catenin.15 P120 catenin is a member of the Armadillo repeat protein family that has multiple isoforms whose expression and localization varies depending on tissues and cell types.15 P120 catenin stabilizes the cadherin–catenin adhesion complex at the plasma membrane, but also has additional roles in the cytoplasm and nucleus, where it has been reported to modulate transcription by associating with the transcriptional repressor Kaiso.25, 26 We therefore examined p120 catenin expression in the Panc1 utilized here and sought to further investigate the effects of p120 catenin re-expression in S2-013 cells on MUC1 modulation of cyclin D1 promoter activity. The western blot from Figure 3a confirms that S2-013 cells do not express p120 catenin and reveal that Panc-1 cells express different isoforms of p120 catenin. Isoforms of p120 catenin differ in amino termini because of alternative splicing and usage of alternate translation-initiating codons, each of which has a distinct start codon in the N-terminal region.27, 28, 29, 30 Isoform 1 contains a coiled-coil domain at the N-terminus; isoform 2 contains the entire ‘regulatory region' isoform 3 has most of regulatory region; isoform 4 lacks the majority of the regulatory region. Panc-1 Neo cells expressed p120 isoform 3 and low levels of isoform 1. Expression of MUC1 in Panc-1 cells stabilized and increased expression of different isoforms of p120 catenin, especially isoforms 1 and 4.


MUC1 regulates cyclin D1 gene expression through p120 catenin and β-catenin.

Liu X, Caffrey TC, Steele MM, Mohr A, Singh PK, Radhakrishnan P, Kelly DL, Wen Y, Hollingsworth MA - Oncogenesis (2014)

P120 catenin was not detected in S2-013 cells and re-expression of different p120 catenin isoforms in this cells showed distinct subcellular localization. (a) Membrane/cytoplasmic extracts and nuclear extracts from S2-013.NEO, S2-013.MUC1F, Panc-1.NEO, Panc-1.MUC1F were subjected to 10% SDS–PAGE and analyzed by immunoblot (IB) with an anti-p120 catenin mAb. The same blot was striped and reprobed with anti-MUC1.CT antibody. (b) Re-expression of different p120 catenin isoforms were confirmed by western blot using the same p120 catenin antibody and MUC1 antibody. (c and d) Immunofluorescence analysis of re-expression of p120 catenin in S2-013 cells. The green color indicates stains for p120 catenin. Blue indicates 4′-6-diamidino-2-phenylindole (DAPI) staining for nuclei. The arrowhead indicates subcellular localization of p120 catenin in the nucleus.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4150213&req=5

fig3: P120 catenin was not detected in S2-013 cells and re-expression of different p120 catenin isoforms in this cells showed distinct subcellular localization. (a) Membrane/cytoplasmic extracts and nuclear extracts from S2-013.NEO, S2-013.MUC1F, Panc-1.NEO, Panc-1.MUC1F were subjected to 10% SDS–PAGE and analyzed by immunoblot (IB) with an anti-p120 catenin mAb. The same blot was striped and reprobed with anti-MUC1.CT antibody. (b) Re-expression of different p120 catenin isoforms were confirmed by western blot using the same p120 catenin antibody and MUC1 antibody. (c and d) Immunofluorescence analysis of re-expression of p120 catenin in S2-013 cells. The green color indicates stains for p120 catenin. Blue indicates 4′-6-diamidino-2-phenylindole (DAPI) staining for nuclei. The arrowhead indicates subcellular localization of p120 catenin in the nucleus.
Mentions: We recently reported that the S2-013 cell line is deficient in expression of p120 catenin, and that re-expression of p120 catenin in these cells stabilized and enhanced steady-state levels of β-catenin.15 P120 catenin is a member of the Armadillo repeat protein family that has multiple isoforms whose expression and localization varies depending on tissues and cell types.15 P120 catenin stabilizes the cadherin–catenin adhesion complex at the plasma membrane, but also has additional roles in the cytoplasm and nucleus, where it has been reported to modulate transcription by associating with the transcriptional repressor Kaiso.25, 26 We therefore examined p120 catenin expression in the Panc1 utilized here and sought to further investigate the effects of p120 catenin re-expression in S2-013 cells on MUC1 modulation of cyclin D1 promoter activity. The western blot from Figure 3a confirms that S2-013 cells do not express p120 catenin and reveal that Panc-1 cells express different isoforms of p120 catenin. Isoforms of p120 catenin differ in amino termini because of alternative splicing and usage of alternate translation-initiating codons, each of which has a distinct start codon in the N-terminal region.27, 28, 29, 30 Isoform 1 contains a coiled-coil domain at the N-terminus; isoform 2 contains the entire ‘regulatory region' isoform 3 has most of regulatory region; isoform 4 lacks the majority of the regulatory region. Panc-1 Neo cells expressed p120 isoform 3 and low levels of isoform 1. Expression of MUC1 in Panc-1 cells stabilized and increased expression of different isoforms of p120 catenin, especially isoforms 1 and 4.

Bottom Line: We observed a significant enhancement in the activation of cyclin D1 promoter-reporter activity in poorly differentiated Panc1.MUC1F cells that overexpress recombinant MUC1 relative to Panc-1.NEO cells, which express very low levels of endogenous MUC1.Further, MUC1 affected subcellular localization of p120 catenin in association with one of the main effectors of P120 catenin, the transcriptional repressor Kaiso, supporting the hypothesis that p120 catenin relieved transcriptional repression by Kaiso.Our data show MUC1 enhances the activities of both β-catenin and p120 catenin.

View Article: PubMed Central - PubMed

Affiliation: Eppley Institute for Research in Cancer and Allied Diseases, University of Nebraska Medical Center, Omaha, NE, USA.

ABSTRACT
MUC1 interacts with β-catenin and p120 catenin to modulate WNT signaling. We investigated the effect of overexpressing MUC1 on the regulation of cyclin D1, a downstream target for the WNT/β-catenin signaling pathway, in two human pancreatic cancer cell lines, Panc-1 and S2-013. We observed a significant enhancement in the activation of cyclin D1 promoter-reporter activity in poorly differentiated Panc1.MUC1F cells that overexpress recombinant MUC1 relative to Panc-1.NEO cells, which express very low levels of endogenous MUC1. In stark contrast, cyclin D1 promoter activity was not affected in moderately differentiated S2-013.MUC1F cells that overexpressed recombinant MUC1 relative to S2-013.NEO cells that expressed low levels of endogenous MUC1. The S2-013 cell line was recently shown to be deficient in p120 catenin. MUC1 is known to interact with P120 catenin. We show here that re-expression of different isoforms of p120 catenin restored cyclin D1 promoter activity. Further, MUC1 affected subcellular localization of p120 catenin in association with one of the main effectors of P120 catenin, the transcriptional repressor Kaiso, supporting the hypothesis that p120 catenin relieved transcriptional repression by Kaiso. Thus, full activation of cyclin D1 promoter activity requires β-catenin activation of TCF-lef and stabilization of specific p120 catenin isoforms to relieve the repression of KAISO. Our data show MUC1 enhances the activities of both β-catenin and p120 catenin.

No MeSH data available.


Related in: MedlinePlus