We had earlier demonstrated that PKC Mu, a novel isozyme of the PKC family of signal transduction proteins, is down regulated in androgen independent (AI) PC. Because PKC Mu is uniquely constitutively membrane bound compared to other members of the PKC family, we explored its interaction with the membrane associated cadherin-catenin complex of proteins, which are involved with tumorogenesis and metastasis. We studied the interaction in androgen dependent (AD) LNCaP prostate cancer cells by Immunofluorescence (IF) and Immunoprecipitaion (IP) assays. In Vivo IF assay was done by culturing cells on glass coverslips for 48h, permeabilized with cold methanol, incubated with PKC Mu (C-20) and E-cadherin (HECD1) or Beta-catenin (6F9) antibodies, treated with FITC-conjugated secondary antibodies and analyzed with a LSM 510 laser scanning microscope. For IP assays, protein lysate was obtained from 80% confluent cells, immunoprecipitated with PKC Mu (C-20) and immunoblotted with E-cadherin (HECD1) or Beta-catenin (6F9) antibodies. Reciprocal IP was done with E-cadherin (HECD1) and immunoblotted with PKC Mu (C-20) antibodies. Confocal microsocopy demonstrated that PKC Mu localized to the cell membrane and cytoplasm in the peri-nuclear areas. PKC Mu co-localized with E-cadherin and Beta-catenin at cell-to-cell junctional areas. Both IP and reciprocal IP experiments confirmed that PKC Mu interacted with E- cadherin and IP assay confirmed that PKC Mu also interacted with Beta-catenin. Because E-cadherins are well known to strongly interact with Beta-catenin, the PKC Mu may interact either directly or indirectly with the members of the cadherin-catenin complex of proteins. Our studies have identified a novel interaction between PKC Mu and cadherin-catenin complex of proteins in vivo and in vitro. The functional significance of PKC Mu in PC is currently unclear. It is conceivable that dysregulation of PKC Mu expression in PC may contribute to PC progression by its interaction with the cadherin-catenin complex of proteins, which are well established markers of cancer progression in other malignancies.
[Proc Amer Assoc Cancer Res, Volume 45, 2004]