Am J Cancer Res 2012;2(5):478-491
Original Article
PAR6B is required for tight junction formation and activated PKCζ localization in
breast cancer
Heather E Cunliffe*, Yuan Jiang*, Kimberly M Fornace, Fan Yang, Paul S Meltzer
Translational Genomics Research Institute (TGen), Phoenix, AZ 85004, USA; Genetics Branch, National Cancer Institute,
National Institutes of Health, Bethesda MD 20892, USA; Royal Veterinary College, University of London, UK. *H.E.C. and Y.J.
contributed equally to this work.
Received June 1, 2012; accepted July 11, 2012; Epub August 20, 2012; Published September 15, 2012
Abstract: Dysregulation of mechanisms that govern the control of epithelial cell polarity, morphology and plasticity are emerging
as key processes in tumor progression. In this study we report amplification and overexpression of PAR6B, an essential
component in epithelial cell tight junction (TJ) formation and maintenance of apico-basal polarity, in breast cancer cell lines.
Analysis of chromosome 20q13.13 in 11 breast cancer cell lines by fluorescence in situ hybridization (FISH) identified a novel
small amplicon centered at PARD6B in 5 cell lines, with copy number ranging from 7 to 27. The presence of the PARD6B
amplicon correlated with PARD6B transcript and PAR6B protein abundance. Expression of related isoforms PARD6A and
PARD6G were detectable at significantly lower levels. PARD6B overexpression correlated with TJ network formation in cultured
cell monolayers. SiRNA-mediated inhibition of PAR6B in MCF7 resulted in loss of TJ assembly and membrane localization of
atypical PKCζ (aPKC), but did not affect adherens junction formation. SiRNA-mediated inhibition of CDC42 in MCF7 also resulted
in loss of TJ networks, confirming the requirement of a complete PAR6-aPKC-CDC42-PAR3 complex to activate and stabilize
TJs. Immunohistochemical analysis of PAR6B expression on breast tumor microarrays indicated exquisite epithelial cell-
specificity. Few quantitative differences in staining were observed between normal epithelium and adjacent tumor margins.
However staining appeared reduced and cytoplasmic in more poorly differentiated tumors. We propose that quantitative
imbalances in the components of pathways governing normal epithelial cell polarity arising from gain or loss of function may
radically alter epithelial cell architecture and contribute to tumor progression. (AJCR0000127).
Keywords: Breast Cancer, DNA amplification, tight junction, siRNA, polarity, adhesion, PARD6B, PAR6B, CDC42, PKCζ
Address all correspondence to:
Dr. Paul Meltzer
Genetics Branch, National Cancer Institute
National Institutes of Health
37 Convent Dr. MSC 4265
Bethesda, MD 20892-4265, USA.
Tel: (301) 496-5266; Fax: (301) 402-3241
E-mail: pmeltzer@mail.nih.gov
Original Article
PAR6B is required for tight junction formation and activated PKCζ localization in
breast cancer
Heather E Cunliffe*, Yuan Jiang*, Kimberly M Fornace, Fan Yang, Paul S Meltzer
Translational Genomics Research Institute (TGen), Phoenix, AZ 85004, USA; Genetics Branch, National Cancer Institute,
National Institutes of Health, Bethesda MD 20892, USA; Royal Veterinary College, University of London, UK. *H.E.C. and Y.J.
contributed equally to this work.
Received June 1, 2012; accepted July 11, 2012; Epub August 20, 2012; Published September 15, 2012
Abstract: Dysregulation of mechanisms that govern the control of epithelial cell polarity, morphology and plasticity are emerging
as key processes in tumor progression. In this study we report amplification and overexpression of PAR6B, an essential
component in epithelial cell tight junction (TJ) formation and maintenance of apico-basal polarity, in breast cancer cell lines.
Analysis of chromosome 20q13.13 in 11 breast cancer cell lines by fluorescence in situ hybridization (FISH) identified a novel
small amplicon centered at PARD6B in 5 cell lines, with copy number ranging from 7 to 27. The presence of the PARD6B
amplicon correlated with PARD6B transcript and PAR6B protein abundance. Expression of related isoforms PARD6A and
PARD6G were detectable at significantly lower levels. PARD6B overexpression correlated with TJ network formation in cultured
cell monolayers. SiRNA-mediated inhibition of PAR6B in MCF7 resulted in loss of TJ assembly and membrane localization of
atypical PKCζ (aPKC), but did not affect adherens junction formation. SiRNA-mediated inhibition of CDC42 in MCF7 also resulted
in loss of TJ networks, confirming the requirement of a complete PAR6-aPKC-CDC42-PAR3 complex to activate and stabilize
TJs. Immunohistochemical analysis of PAR6B expression on breast tumor microarrays indicated exquisite epithelial cell-
specificity. Few quantitative differences in staining were observed between normal epithelium and adjacent tumor margins.
However staining appeared reduced and cytoplasmic in more poorly differentiated tumors. We propose that quantitative
imbalances in the components of pathways governing normal epithelial cell polarity arising from gain or loss of function may
radically alter epithelial cell architecture and contribute to tumor progression. (AJCR0000127).
Keywords: Breast Cancer, DNA amplification, tight junction, siRNA, polarity, adhesion, PARD6B, PAR6B, CDC42, PKCζ
Address all correspondence to:
Dr. Paul Meltzer
Genetics Branch, National Cancer Institute
National Institutes of Health
37 Convent Dr. MSC 4265
Bethesda, MD 20892-4265, USA.
Tel: (301) 496-5266; Fax: (301) 402-3241
E-mail: pmeltzer@mail.nih.gov
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American Journal of Cancer Research
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