Activation of protein kinase (PK) C has been considered cancer-promoting based on early data showing that phorbol esters could induce oncogenes is through the activation of so called classic and novel PKC isozymes. Although many human cancers exhibit increased levels of specific PKC isozymes there are conflicting data which show that specific isozymes can act as either promoters or suppressors of oncogenesis depending on tissue context. In human prostate cancer (CaP), PKC, levels increase with malignancy whereas PKC* levels are constant, yet only the overexpression of PKC, is sufficient to induce prostatic intraepithelial neoplasia (PIN). Therefore, characterization of the pro-oncogenic PKC-induced signaling pathways and mediators is paramount to our overall understanding of CaP disease progression. Our lab identified a novel PKC substrate, SSeCKS/Gravin/AKAP12 (""""""""SSeCKS"""""""") that is also a scaffold for both PKC and PKA, capable of attenuating PKC kinase activity and altering its cellular compartmentalization, while regulating PKA through compartmentalization only. SSeCKS displays many of the hallmarks of a tumor suppressor in prostate cancer: it is severely downregulated in human CaP cell lines and tissues with Gleason sums =6, SSeCKS reexpression suppresses macroscopic CaP metastasis growth by inhibiting VEGF-induced neovascularization, genetic knockout (KO) in mice induces prostatic hyperplasia and focal dysplasia with evidence of epithelial cell senescence, and KO mouse fibroblasts (MEF) display premature senescence correlating with polyploidy and binucleation. Premature senescence seems Rb-dependent because it can be suppressed by HPV-16E7orpRb-siRNA but not byHPV-16 E6 or p53-siRNA. In keeping with SSeCKS'scaffolding function for PKC, KO-MEF have >2-fold higher total PKC activity than WT-MEF, with a >3-fold increase in PKC* activity. Based on preliminary data from expression microarray and biochemical experiments, we hypothesize the loss of SSeCKS induces hyperactive PKC*/,,which in turn induce i) cytokinesis defects through the activation of RhoA and LIMK, and through the suppression of the mitotic exit network kinase, WARTS, and ii) Rb-dependent senescence by increasing reactive oxygen species (ROS) mediators such as p47phox. We believe SSeCKS controls cytokinesis directly because a pool of SSeCKS enriches in the cleavage furrow during telophase where PKC* and , are known to normally regulate abscission by coordinating the timing of RhoA and LIMK activation, and thus, formation and contraction of the actomyosin abscission ring. Our overall aim is to use genetic, biochemical, and fluorescence and time-lapse microscopy techniques to dissect this novel SSeCKS-PKC pathway in WT vs. KO MEF and murine prostate epithelial cells (PrEC), and in human PrEC vs. CaP cell lines, using readouts of senescence/ polyploidy/ binucleation and cytokinesis completion (Aim 1), podosome formation and tumor invasiveness (Aim 2), and CaP formation, invasiveness and metastatic potential in transgenic mouse models (Aim 3). The experiments proposed are envisioned to elucidate the mechanism and pathways controlled by the SSeCKS/PKC scaffold complex and how dysregulation by the loss of SSeCKS contributes to CaP progression.

Public Health Relevance

Our project focuses on SSeCKS, a protein that seems to suppress prostate cancer progression especially metastasis, by binding two opposing signaling proteins, PKA and PKC, and controlling their function by regulating when and where in the cell they are activated. We now propose to elaborate on pro-cancer pathways that PKC induces, yet which are normally regulated by SSeCKS. These studies will elucidate how the loss of SSeCKS expression in prostate cancer increases disease progression by allowing PKC to be hyperactivated and dysregulated.

National Institute of Health (NIH)
National Cancer Institute (NCI)
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Tumor Progression and Metastasis Study Section (TPM)
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Ault, Grace S
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Roswell Park Cancer Institute Corp
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Muramatsu, Masashi; Akakura, Shin; Gao, Lingqiu et al. (2018) SSeCKS/Akap12 suppresses metastatic melanoma lung colonization by attenuating Src-mediated pre-metastatic niche crosstalk. Oncotarget 9:33515-33527
Muramatsu, Masashi; Gao, Lingqiu; Peresie, Jennifer et al. (2017) SSeCKS/AKAP12 scaffolding functions suppress B16F10-induced peritoneal metastasis by attenuating CXCL9/10 secretion by resident fibroblasts. Oncotarget 8:70281-70298
Chattopadhyay, Indranil; Wang, Jianmin; Qin, Maochun et al. (2017) Src promotes castration-recurrent prostate cancer through androgen receptor-dependent canonical and non-canonical transcriptional signatures. Oncotarget 8:10324-10347
Dwyer, Sheila Figel; Gao, Lingqiu; Gelman, Irwin H (2015) Identification of novel focal adhesion kinase substrates: role for FAK in NF?B signaling. Int J Biol Sci 11:404-10
Gelman, Irwin H; Peresie, Jennifer; Eng, Kevin H et al. (2014) Differential requirement for Src family tyrosine kinases in the initiation, progression, and metastasis of prostate cancer. Mol Cancer Res 12:1470-9
Dwyer, Sheila Figel; Gelman, Irwin H (2014) Cross-Phosphorylation and Interaction between Src/FAK and MAPKAP5/PRAK in Early Focal Adhesions Controls Cell Motility. J Cancer Biol Res 2:
Ko, Hyun-Kyung; Akakura, Shin; Peresie, Jennifer et al. (2014) A transgenic mouse model for early prostate metastasis to lymph nodes. Cancer Res 74:945-53
Ko, Hyun-Kyung; Guo, Li-wu; Su, Bing et al. (2014) Suppression of chemotaxis by SSeCKS via scaffolding of phosphoinositol phosphates and the recruitment of the Cdc42 GEF, Frabin, to the leading edge. PLoS One 9:e111534
Cha, Jong-Ho; Wee, Hee-Jun; Seo, Ji Hae et al. (2014) Prompt meningeal reconstruction mediated by oxygen-sensitive AKAP12 scaffolding protein after central nervous system injury. Nat Commun 5:4952
Smeets, Bart; Boor, Peter; Dijkman, Henry et al. (2013) Proximal tubular cells contain a phenotypically distinct, scattered cell population involved in tubular regeneration. J Pathol 229:645-59

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