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Name Christopher Apostolatos
Organization or Institution University of South Florida
Presentation Type Poster
Topic Biochemistry / Chem Bio.
Title

C-Jun and FOXO1 play key roles in the overexpression of oncogenic PKC-ἱ in human prostate and melanoma cell lines

Author(s)

Wishrawana Sarathi Ratnayake, Christopher Apostolatos, André Apostolatos and Mildred Acevedo-Duncan

Author Institution(s)

University of South Florida

Abstract

Protein Kinase C-ἱ (PKC-ἱ) is an anti-apoptotic oncogene overexpressed in multiple cancers including prostate, melanoma, ovarian, and breast glioma. We have previously shown that PKC-ἱ is part of a cycle that helps cancer cell avoid senescence by releasing the transcription factor NF-kB and promoting apoptotic resistance while promoting epithelial-mesenchymal transition in melanoma {Ratnayake, et al. Int. J. Oncol. 51(5), (2017), 1370-1382}. PKC-ἱ is activated externally by factors like loss of PTEN and TGFβ stimulation. However, while under the inhibition of PKC-ἱ specific inhibitors, Drop of PKC-ι expression along with phosphorylated levels are markedly reduced, suggesting PKC-ἱ plays a role in regulating its own expression. A previous study showed the ELK1 transcription factor to be a regulator of PKC-ἱ (Put the complete citation as above for Gustafson 2003). In this study, transcription factors such as C-Jun, ISGF3, PAX3, EGR1, and FOXO1 which bind on or near the promoter sequence of the PRKCI gene and their role in PKC-ἱ regulation was analyzed mainly in prostate cancer cell lines (DU145 and PC3) and melanoma cell lines (SK-MEL-2 and MeWo). Each transcription factor was systematically silenced. Western blots revealed that expression of PKC-ἱ significantly altered in C-jun and FOXO1 treated cells showing C-Jun followed by FOXO1 are the two major transcription factors which involve in PKC-ι expression in prostate cells compared to FOXO1 in melanoma cells. This suggests that these transcription factors act as activators that regulate PKC-iota expression and can be downstream targets of PKC-iota. We found that regulation of PKC-ι expression was governed mainly through NF-κB pathway. siRNA knockdown of NF-κB p65 and application of NF-κB inhibitors  confirmed the above observation. We have also used immuno-paired-antibody detection assay was used to determine which pathways were affected with PKC-ι inhibitor treatments other than NF-κB pathway. qPCR and microarray were performed to analyze the transcriptome of treated cells to match protein levels with mRNA levels.  We have analyzed multiple targets both up and downstream of PKC-ἱ to determine the mechanism of PKC-ἱ self-regulation. Overall results suggest that PKC-ι inhibition leads to downregulates its own expression therefore can be used as a novel therapeutic target in prostate and melanoma cancers.