ABSTRACT
Nuclear hormone receptor family member PPARγ plays an important role in mammary gland tumorigenesis. Previous studies have shown PPARγ has cytoplasmic activities upon tetradecanoyl phorbol acetate (TPA) stimulation. However, the clinical pathological significance of cytoplasmic PPARγ is not completely understood in human breast cancer. Skp2 is oncogenic, and its frequent amplification and overexpression correlated with the grade of malignancy. In this study, the role of cytoplasmic PPARγ and Skp2 expression was investigated in human breast cancer progression. Therefore, immunohistochemical analysis was performed on formalin-fixed paraffin sections of 70 specimens. Furthermore, Western blot and immunofluorescence microscopy analysis were used to study the relationship between expression of cytoplasmic PPARγ and Skp2 expression in human breast cancer cells in vitro. Results showed that the expression of cytoplasmic PPARγ was positively correlated with Skp2 expression (p < 0.05), and correlated significantly with estrogen receptor (p = 0.026) and pathological grade (p = 0.029), respectively. In addition, Skp2 overexpression can provoke cytoplasmic localization of PPARγ upon MEK1-dependent mechanisms in human breast cancer cells by nuclear-cytosolic fractionation technology and immunofluorescence microscopy analysis. Using RNA interference technology, we also found that down-regulated Skp2 reduced the phosphorylation level of MEK1 and significantly reversed TPA-induced nuclear export of PPARγ in MDA-MB-231 cells. The changes in the subcellular localization of PPARγ may represent a novel target for selective interference in patients with breast cancer.
Subject(s)
Breast Neoplasms/metabolism , MAP Kinase Kinase 1/metabolism , PPAR gamma/metabolism , S-Phase Kinase-Associated Proteins/metabolism , Breast Neoplasms/genetics , Butadienes/pharmacology , Cell Line, Tumor , Enzyme Inhibitors/pharmacology , Female , Flavonoids/pharmacology , Humans , MAP Kinase Kinase 1/antagonists & inhibitors , MAP Kinase Signaling System , MCF-7 Cells , Nitriles/pharmacology , PPAR gamma/biosynthesis , PPAR gamma/genetics , Phosphorylation/genetics , Protein Kinase Inhibitors/pharmacology , RNA Interference , RNA, Small Interfering , Receptors, Estrogen/metabolism , S-Phase Kinase-Associated Proteins/biosynthesis , S-Phase Kinase-Associated Proteins/genetics , Tetradecanoylphorbol Acetate/pharmacologyABSTRACT
Members of tumour necrosis factor (TNF) family usually trigger both survival and apoptotic signals in various cell types. Heat shock proteins (HSPs) are conserved proteins implicated in protection of cells from stress stimuli. However, the mechanisms of HSPs in TNFalpha-induced signalling pathway have not been fully elucidated. We report here that HSP70 over-expression in human colon cancer cells can inhibit TNFalpha-induced NFkappaB activation but promote TNFalpha-induced activation of c-Jun N-terminal kinase (JNK) through interaction with TNF receptor (TNFR)-associated factor 2 (TRAF2). We provide evidence that HSP70 over-expression can sequester TRAF2 in detergent-soluble fractions possibly through interacting with TRAF2, leading to reduced recruitment of receptor-interacting protein (RIP1) and IkappaB alpha kinase (IKK) signalosome to the TNFR1-TRADD complex and inhibited NFkappaB activation after TNFalpha stimuli. In addition, we found that HSP70-TRAF2 interaction can promote TNFalpha-induced JNK activation. Therefore, our study suggests that HSP70 may differentially regulate TNFalpha-induced activation of NFkappaB and JNK through interaction with TRAF2, contributing to the pro-apoptotic roles of HSP70 in TNFalpha-induced apoptosis of human colon cancer cells.
Subject(s)
HSP70 Heat-Shock Proteins/metabolism , JNK Mitogen-Activated Protein Kinases/metabolism , NF-kappa B/metabolism , TNF Receptor-Associated Factor 2/metabolism , Tumor Necrosis Factor-alpha/pharmacology , Apoptosis/drug effects , Blotting, Western , Cell Line , Cell Line, Tumor , Enzyme Activation , HSP70 Heat-Shock Proteins/genetics , HT29 Cells , Humans , Membrane Microdomains/metabolism , Protein Binding , RNA Interference , Receptors, Tumor Necrosis Factor, Type I/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Signal Transduction , TNF Receptor-Associated Death Domain Protein/metabolism , TNF Receptor-Associated Factor 2/genetics , TransfectionABSTRACT
To explore the real association between p53 codon 72 polymorphism and lung cancer risk, a pooled analysis of 32 case-control studies involving 19,255 subjects was conducted. When all 32 studies were pooled into the analysis, significantly elevated lung cancer risks were associated with variant genotypes in all genetic models (for Pro/Arg vs. Arg/Arg: OR 1.21, 95% CI 1.01-1.23; for Pro/Pro vs. Arg/Arg: OR 1.20, 95% CI 1.03-1.39; for Pro/Pro + Pro/Arg vs. Arg/Arg: OR 1.14, 95% CI 1.03-1.25; for Pro/Pro vs. Arg/Arg + Pro/Arg: OR 1.06, 95% CI 1.01-1.12). In the subgroup analysis by ethnicity, histological type, or smoking status, significantly increased risks were found in subgroups such as Asians, Caucasians, lung adenocarcinoma patients, or smokers, respectively. In conclusion, our results suggest that the Pro allele at p53 codon 72 is emerging as a low-penetrance susceptibility allele for lung cancer development.
