Abstract 2857: Vimentin is a novel PKC-ι target facilitating migration and invasion in melanoma

Abstract

Abstract Vimentin plays an important role in gaining rear-to-front polarity for mesenchymal cells, making it a hallmark of cancer metastasis. Epithelial-mesenchymal transition (EMT) is a vital process in tumor progression that converts stationary epithelial cells into motile mesenchymal phenotype by downregulating E-cadherin and upregulating Vimentin expression. Molecular dynamics of Vimentin intermediate filaments (VIFs) play several key roles in cancer metastasis. Vimentin is activated via phosphorylation through various kinases. Recently, we reported that Protein Kinase C-iota (PKC-ι), activates Vimentin in melanoma while inducing EMT. We also reported the effects of PKC-ι inhibition on mitigating EMT in melanoma based on PKC- ι specific inhibitors {Ratnayake, W.S., et al. Int. J. Oncol. 51(5), (2017), 1370-1382 https://doi.org/10.3892/ijo.2017.4131 and Ratnayake, W.S., et al. Cell Adhes. Migr. (2018) https://doi.org/10.1080/19336918.2018.1471323}. This novel finding is very important with respect to cancer progression since PKC-ι is a well-established oncogene in various tumor types. Here, we analyzed the VIF dynamic changes upon PKC-ι inhibition using specific inhibitors targeting multiple activation sites on Vimentin. In addition to siRNA knockdown of expression of PKC-ι, we also report the effects of siRNA knockdown of two transcription factors, Paired related homeobox1 (PRRX1) and Zinc finger protein SNAI1 (SNAIL1) on the expressions of Vimentin, PKC-ι and E-cadherin. Herein we show that inhibition of PKC-ι downregulate the transcriptional activities of PRRX1 and SNAIL1, which was demonstrated with diminished levels of Vimentin and elevated levels of E-cadherin. Western blot, confocal immunofluorescence and immune-gold electron microscopic results suggested that PKC-ι targets multiple activation sites (S33, S39 and S56) on Vimentin without having an effect on S6 and S71 phosphorylation sites. Results also suggest that PKC-ι inhibition leads to diminution of activities of PRRX1 and SNAIL1 thereby further downregulation of Vimentin while upregulating E-cadherin in order to continue retardation of EMT in-vitro. Overall results indicate that PKC-ι is essential for VIF dynamics regulation during the metastasis in-vitro and therefore can be used as a novel biomarker to assess and mitigate melanoma metastasis. Citation Format: Wishrawana Sarathi Ratnayake, Sloan Breedy, Christopher Apostolatos, Robert Hill, Clare Dennison, Mildred Acevedo-Duncan. Vimentin is a novel PKC-ι target facilitating migration and invasion in melanoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 2857.