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KLF5 promotes apoptosis induced by phorbol ester as an effector of the autocrine factor TNFα in LNCaP prostate cancer cells.

ABSTRACT: Krüppel-like factor 5 (KLF5) is frequently deleted and inactivated in prostate cancer, and exerts tumor-suppressing function in prostate cancer cells. However, the function of KLF5 in the apoptosis of prostate cancer cells remains unclear. In the present study, the effect of KLF5 on phorbol 12-myristate 13-acetate (PMA)-induced apoptosis was investigated in prostate cancer LNCaP cells. It was demonstrated that PMA induced the expression of KLF5 at the mRNA and protein level. To identify whether KLF5 regulates the activity of the downstream pathway, stable KLF5 knockdown or overexpression cell lines were constructed with lentivirus harboring shRNA targeting KLF5 or full-length KLF5 in LNCaP cells. Knockdown of KLF5 significantly decreased PMA-induced apoptosis, while cell apoptosis was significantly increased following KLF5 overexpression compared with the corresponding control groups. Consistently, expression of cleaved poly(ADP-ribose) polymerase and caspase-3 induced by PMA was decreased following KLF5 knockdown and increased following KLF5 overexpression. Using the control medium from cells treated with PMA, it was demonstrated that KLF5 is required for the control medium to induce apoptosis. c-Jun N-terminal kinase (JNK) activity is essential for the apoptosis induced by PMA. It was revealed that knockdown of KLF5 decreased, while overexpression of KLF5 increased the phosphorylation of JNK induced by PMA and control medium treatment. Furthermore, inhibition of tumor necrosis factor α (TNFα) decreased KLF5 expression and significantly decreased cell apoptosis induced by PMA, and control medium. This data indicates that KLF5 is essential for the apoptosis induced by PMA in LNCaP prostate cancer cells. Furthermore, KLF5 is essential for activity of the autocrine factor TNFα, which is secreted by cells treated with PMA and mediates the function of PMA-induced apoptosis through regulating the activity of JNK signaling pathway. These results provide novel insights into the complexity of the signaling pathways regulating apoptosis in prostate cancer cells, which could aid in the development of novel treatments for patients with prostate cancer.

SUBMITTER: Shi Q

PROVIDER: S-EPMC5529878 | biostudies-other | 2017 Aug

REPOSITORIES: biostudies-other

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Project description:BACKGROUND: Malignancy alters cellular complex lipid metabolism and membrane lipid composition and turnover. Here, we investigated whether tumorigenesis in cancer-derived prostate epithelial cell lines influences protein kinase C-linked turnover of ethanolamine phosphoglycerides (EtnPGs) and alters the pattern of ethanolamine (Etn) metabolites released to the medium. METHODS: Prostate epithelial cell lines P4E6, LNCaP and PC3 were models of prostate cancer (PCa). PNT2C2 and PNT1A were models of benign prostate epithelia. Cellular EtnPGs were labelled with [1-(3)H]-Etn hydrochloride. PKC was activated with phorbol ester (TPA) and inhibited with Ro31-8220 and GF109203X. D609 was used to inhibit PLD (phospholipase D). [(3)H]-labelled Etn metabolites were resolved by ion-exchange chromatography. Sodium oleate and mastoparan were tested as activators of PLD2. Phospholipase D activity was measured by a transphosphatidylation reaction. Cells were treated with ionomycin to raise intracellular Ca(2+) levels. RESULTS: Unstimulated cell lines release mainly Etn and glycerylphosphorylEtn (GPEtn) to the medium. Phorbol ester treatment over 3h increased Etn metabolite release from the metastatic PC3 cell line and the benign cell lines PNT2C2 and PNT1A but not from the tumour-derived cell lines P4E6 and LNCaP; this effect was blocked by Ro31-8220 and GF109203X as well as by D609, which inhibited PLD in a transphosphatidylation reaction. Only metastatic PC3 cells specifically upregulated Etn release in response to TPA treatment. Oleate and mastoparan increased GPEtn release from all cell lines at the expense of Etn. Ionomycin stimulated GPEtn release from benign PNT2C2 cells but not from cancer-derived cell lines P4E6 or PC3. Ethanolamine did not stimulate the proliferation of LNCaP or PC3 cell lines but decreased the uptake of choline (Cho). CONCLUSIONS: Only the metastatic basal PC3 cell line specifically increased the release of Etn on TPA treatment most probably by PKC activation of PLD1 and increased turnover of EtnPGs. The phosphatidic acid formed will maintain a cancer phenotype through the regulation of mTOR. Ethanolamine released from cells may reduce Cho uptake, regulating the membrane PtdEtn:PtdCho ratio and influencing the action of PtdEtn-binding proteins such as RKIP and the anti-apoptotic hPEBP4. The work highlights a difference between LNCaP cells used as a model of androgen-dependent early stage PCa and androgen-independent PC3 cells used to model later refractory stage disease.

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KLF5 promotes apoptosis induced by phorbol ester as an effector of the autocrine factor TNFα in LNCaP prostate cancer cells.

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