Expression Of SET8 Research Articles

SET8 inhibition preserves PTEN to attenuate kidney cell apoptosis in cisplatin nephrotoxicity.

The aberrant expression of SET8, a histone methyltransferase that mediates H4 lysine 20 mono-methylation (H4K20me1), is implicated in the pathogenesis of various tumors, however, its role in acute kidney injury (AKI) is unknown. Here we showed that SET8 and H4K20me1 were upregulated in the murine kidney with AKI induced by cisplatin, along with increased renal tubular cell injury and apoptosis and decreased expression of E-cadherin and Phosphatase and Tensin Homolog (PTEN). Suppression of SET8 by UNC0379 improved renal function, attenuated tubule damage, and restored expression of PTEN, but not E-cadherin. UNC0379 was also effective in lessening cisplatin-induced DNA damage response (DDR) as indicated by reduced expression of γ-H2AX, p53, p21, and alleviating cisplatin-impaired autophagy as shown by retained expression of Atg5, Beclin-1, and CHMP2A and enhanced levels of LC3-II in the kidney. Consistently, inhibition of SET8 with either UNC0379 or siRNA mitigated apoptosis and DDR, and restored autophagy, along with PTEN preservation in cultured renal proximal tubular epithelial cell (TKPTs) exposed to cisplatin. Further studies showed that inhibition of PTEN with Bpv or siRNA potentiated cisplatin-induced apoptosis, DDR, and hindered autophagy, and conversely, alleviated by overexpression of PTEN in TKPTs. Finally, blocking PTEN largely abolished the inhibitory effect of UNC0379 on apoptosis. Taken together, these results suggest that SET8 inhibition protects against cisplatin-induced AKI and renal cell apoptosis through a mechanism associated with the preservation of PTEN, which in turn inhibits DDR and restores autophagy.

SET8 Inhibition Potentiates Radiotherapy by Suppressing DNA Damage Repair in Carcinomas

ObjectiveSET8 is a member of the SET domain-containing family and the only known lysine methyltransferase (KMT) that monomethylates lysine 20 of histone H4 (H4K20me1). SET8 has been implicated in many essential cellular processes, including cell cycle regulation, DNA replication, DNA damage response, and carcinogenesis. There is no conclusive evidence, however, regarding the effect of SET8 on radiotherapy. In the current study we determined the efficacy of SET8 inhibition on radiotherapy of tumors and the underlying mechanism. MethodsFirst, we explored the radiotherapy benefit of the SET8 expression signature by analyzing clinical data. Then, we measured a series of biological endpoints, including the xenograft tumor growth in mice and apoptosis, frequency of micronuclei, and foci of 53BP1 and γ-H2AX in cells to detect the SET8 effects on radiosensitivity. RNA sequencing and subsequent experiments were exploited to verify the mechanism underlying the SET8 effects on radiotherapy. ResultsLow expression of SET8 predicted a better benefit to radiotherapy in lung adenocarcinoma (LUAD) and invasive breast carcinoma (BRCA) patients. Furthermore, genetic deletion of SET8 significantly enhanced radiation treatment efficacy in a murine tumor model, and A549 and MCF7 cells; SET8 overexpression decreased the radiosensitivity. SET8 inhibition induced more apoptosis, the frequency of micronuclei, and blocked the kinetics process of DNA damage repair as 53BP1 and γ-H2AX foci remained in cells. Moreover, RNF8 was positively correlated with the SET8 impact on DNA damage repair. ConclusionOur results demonstrated that SET8 inhibition enhanced radiosensitivity by suppressing DNA damage repair, thus suggesting that SET8 potentiated radiotherapy of carcinomas. As new inhibitors of SET8 are synthesized and tested in preclinical and clinical settings, combining SET8 inhibitors with radiation warrants consideration for precise radiotherapy.

Fasting Induces Hepatocellular Carcinoma Cell Apoptosis by Inhibiting SET8 Expression.

Background Hepatocellular carcinoma (HCC) is a life-threatening cancer, and the Kelch-like ECH-associated protein 1 (Keap1)/NF-E2-related factor 2 (Nrf2)/antioxidant response element (ARE) signalling pathway plays a crucial role in apoptosis resistance in cancer cells. Fasting is reported to mediate tumour growth reduction and apoptosis. SET8 is involved in cancer proliferation, invasiveness, and migration. However, whether SET8 participates in fasting-mediated apoptosis in HCC remains unclear. Methods We used immunohistochemical staining to analyse the expression of SET8, Keap1, and Nrf2 in HCC tissues. Cell viability, apoptosis, and cellular reactive oxygen species (ROS) were assessed, and Western blot and qPCR analyses were used to examine the expression of Keap1/Nrf2 in HCC cells under fasting, SET8 overexpression, and PGC1α overexpression conditions. Mass spectrometry, coimmunoprecipitation, and confocal microscopy were used to determine whether PGC1α overexpression conditions. Mass spectrometry, coimmunoprecipitation, and confocal microscopy were used to determine whether PGC1In vivo experiments were performed to verify the conclusions from the in vitro experiments. Results Our data indicate that SET8 expression is associated with poor survival in HCC patients. Both in vitro experiments. in vivo experiments were performed to verify the conclusions from the α overexpression conditions. Mass spectrometry, coimmunoprecipitation, and confocal microscopy were used to determine whether PGC1α overexpression conditions. Mass spectrometry, coimmunoprecipitation, and confocal microscopy were used to determine whether PGC1Conclusions The results of our study demonstrate that fasting induces HCC apoptosis by inhibiting SET8 expression and that SET8 interacts with PGC1α to activate the Nrf2/ARE signalling pathway by inhibiting Keap1 expression.α overexpression conditions. Mass spectrometry, coimmunoprecipitation, and confocal microscopy were used to determine whether PGC1

Abstract 1389: Effects of geinstein on histone modifying enzyme expression in prostate cancer

Abstract The present study sought to examine the expression of two histone modifying enzymes, SET8 and SIRT1, in human prostate cancer cell model when exposed to increasing concentrations of genistein. Genistein is a phytoestrogen found mainly in soy-based foods and is of importance due to the varying intake levels worldwide. This compound is thought to have chemopreventive properties and it is hypothesized to exert its affects by epigenetically modifying chromatin organization within cells. As an estrogen-like compound, genistein may exert effects similar to that of estrogen. For this reason, SET8, a histone methyltransferase, and, SIRT1, a histone deacetylase, were studied. Previously, these histone modifying enzymes were found to play a role in estrogen signaling. In the present study, cells were treated either for 24 hours (acute) or 5 days (chronic), total RNA was extracted, followed by an RT-PCR, and amplification of the genes of interest via PCR. Changes in gene expression were quantitated using ImageJ. Overall, it was concluded that the exposure to genistein impacts the gene expression of both SET8 and SIRT1 within human prostate cancer cells. Note: This abstract was not presented at the meeting. Citation Format: Lanni Aquila, Nick Smothermon, Lisa Morey. Effects of geinstein on histone modifying enzyme expression in prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 1389. doi:10.1158/1538-7445.AM2017-1389

Expression of SET8 in renal cell carcinoma and the effects of SET8 silencing on proliferation and migration of 786-O cells

Objective To investigate the expression of lysine methyltransferase SET8 protein in the clear cell carcinoma of kidney and the effects of silencing SET8 gene by shRNA on proliferation and migration of human renal cell carcinoma cell line 786-O in vitro. Methods Immunohistochemistry(IHC) was employed to detect the expression of SET8 in 156 tissue samples of clear cell renal cell carcinoma in the study between December, 2006 and December, 2011.Four pairs of SET8-shRNA were designed and synthesized. Transfection was performed with cationic lipid vectors(LipofectamineTM2000). The transfection efficiency was observed by fluorescence confocal microscopy. The effecive shRNA sequences were screened by Western blot. After transfection with effective shRNA sequences, 786-O cells were divided into 3 groups: (1)SET8-shRNA transfection; (2)negative control transfection; (3)blank control.The assays of methyl thiazolyl tetrazolium(MTT), colony formation and scratch wound were applied to examine variations in cell proliferation, colony formation and migration. Results The positive rate of SET8 expression was 98.7%(154/156), and its expression was closely associated with TNM staging, the size of tumor and lymph node metastasis(all P<0.05). The transfection efficiency was around 70% with a concentration of SET8-shRNA 2μg/L and lipofectamine 2000 4μl/well. SET8-shRNA2 was chosen as the effective sequences with a suppression ratio up to 64%. At 48h post-transfection, the expression of SET8 protein was significantly lower in SET8-shRNA2 group than that in negative control transfection group and blank control group(t=-38.174, P<0.01; t=-58.766, P<0.01). While SET8 gene was silenced by shRNA in SET8-shRNA transfection group, the proliferation, colony formation and migration ability were decreased markedly. Conclusion SET8 protein plays a significant role in the carcinogenesis and progression of renal cell carcinoma. SET8-shRNA can effectively inhibit the expression of SET8 protein in 786-O cells and thus suppress malignant phenotypes of RCC. Key words: Lysine methyltransferaseSET8; Clear cell renal cell carcinoma; 786-O cell; Cell proliferation; Colony formation; Migration

Polymorphism at the miR-502 binding site in the 3' untranslated region of SET8 gene is associated with the risk of clear cell renal cell carcinoma

To investigate the relationship between single nucleotide polymorphism of SET8 gene and the risk of clear cell renal cell carcinoma (CCRCC). We selected 140 CCRCC patients and 130 healthy controls in this case-control study.Genotype of single nucleotide polymorphism (rs16917496) at the miR-502 binding site in the 3'UTR of SET8 mRNA in the CCRCC patients and healthy controls was tested and the association between genotype and risk of cancer was assessed. The expression of SET8 was determined by immunohistochemistry and the relationship between expression of SET8 and genotype of rs16917496 was analyzed. In the control group, CC, CT and TT genotypes were found in 30, 32 and 68 persons, respectively, while in the CCRCC patients, CC, CT and TT genotypes were found in 14 , 47 and 79 cases, respectively.The frequencies of rs16917496 CT and TT genotypes in the CCRCC group were significantly higher than those in the control group (P<0.05). Compared with the CC genotype, patients with CT and TT genotypes were more susceptible to develop CCRCC (P<0.05). CT and TT genotypes of rs16917496 at the miR-502 binding site of the SET8 gene were associated with expression of SET8. Genotype of the SNP rs16917496 at the miR-502 binding site in the 3' untranslated region of the SET8 gene is associated with the expression of SET8 protein. Analysis of genetic polymorphisms in miRNA binding sites may help to identify the subgroups of population susceptible to CCRCC.

Differential Effects of Estradiol and Bisphenol A on SET8 and SIRT1 Expression in Ovarian Cancer Cells.

Exposure to estrogenic compounds has been shown to epigenetically reprogram the female reproductive tract and may contribute to ovarian cancer. The goal of this study was to compare the effect of estradiol or bisphenol A (BPA) on the expression of histone-modifying enzymes (HMEs) in ovarian cancer cells. Using 2 human ovarian cancer cell lines, we examined the expression of SET8, a histone methyltransferase, and SIRT1, a histone deacetylase, after exposure to estrogen or BPA. These experiments were carried out in complete media (fetal bovine serum) that contain natural hormones to understand the impact of additional exposure to estrogen or BPA on HME expression. We found differential expression of the HMEs in the different models examined and between the different compounds. Further, we determined that the changes in gene expression occurred via estrogen receptor signaling using the estrogen receptor antagonist, ICI 182,780 (fulvestrant).

MiR-502/SET8 regulatory circuit in pathobiology of breast cancer

Our previous research and extensive epidemiological studies reproducibly demonstrated that miR-502 potentially targeted the expression of H4K20 methyltransferase SET8 in a wide spectrum of cancer. Yet, the direct targeting of SET8 by miR-502 has not been definitively proven. The clinical significance of the miR-502/SET8 regulatory circuit is also not clear. Here, we conducted cell-based experiments and clinical studies in a cohort of 279 breast cancer samples. We provide evidence that SET8 is a direct target of miR-502. Treatment with miR-502 or downregulation of SET8 suppressed cell proliferation and cell cycle, and reduced cell migration, invasion and EMT. Clinical analyses showed the miR-502 expression was lower in tumor tissues than in adjacent non-tumor tissues and had a significant inverse correlation with that of SET8. Furthermore, high expression of SET8 was significantly associated with poor overall survival (OS) and disease free survival (DFS) of breast cancer. The low expression ratio of miR-502 to SET8 mRNA was also significantly associated with poor OS. Thus, the miR-502/SET8 regulatory circuit emerges as a key regulator of the pathobiology of cancer and a focal point for possible therapeutic intervention.

AB252. The positive expression of SET8 possesses prognostic relevance and mediates Wnt target gene activation in renal cell carcinoma

BackgroundRenal cell carcinoma (RCC) is common renal malignancy within poor prognosis. Histone methylase SET8 plays vital roles in tumorigenesis through regulating Wnt/β-catenin signaling and other cancer-related pathways. Here, we studied the expression and function of SET8 inccRCCMethodsOur study was performed immunohistochemistry to assess the overall levels of SET8 and β-catenin in a tissue microarray with 50 RCC specimens. Then, we examined the protein levels of SET8 in RCC and reduced the expression of SET8 by shRNA-mediated partial depletion in order to systematically investigate whether SET8 was involved inRCC carcinogenesis and tumor development measured by 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide (MTT) assay, trans well assay and flow cytometry, respectively. Protein and mRNA expression were assayed by western blotting and quantitative real-time PCR (qPCR). The association between proteins was assayed by co-immunoprecipitation.ResultsSET8 levels and the abnormal expression of β-catenin in the diverse RCC subtypes were inversely correlated with Fuhrman grading, pT stage, and metastasis. Further, shRNA-mediated partial depletion of SET8 reduced 786-0 and ACHN cell viability, arrested them at S phase of the cell cycle and inhibited their abilities of invasion and metastasis. Moreover, western blot implied that several protein expressions of Wnt target genes were inhibited by SET8 interference. It was found that SET8 directly interacted with lymphoid enhancing factor-1 (LEF1)/TCF4, and regulated the transcription of Wnt-activated genes, possibly through H4K20monomethylation at the target gene promoters.ConclusionsCollectively, these results suggested that SET8 could serve as a promising biomarker and contribute to predicting distal metastasis and prognosis of patient with RCC.

Differential effect of estradiol and bisphenol A on Set8 and Sirt1 expression in prostate cancer

Exposure to estrogenic compounds has been shown to epigenetically reprogram the prostate and may contribute to prostate cancer. The goal of this study was to determine the effect of physiological doses of estradiol and bisphenol A (BPA) on the expression of histone modifying enzymes (HMEs) in prostate cancer. Using two human prostate cancer cell lines we examined the expression of Set8, a histone methyltransferase, and Sirt1, a histone deacetylase, after exposure to estrogen or BPA. These experiments were carried out in the presence of natural hormones to understand the impact of additional exposure to estrogen or BPA on HME expression. We found differential expression of the HMEs in the different models and between the different compounds. Further, we determined that the changes in gene expression occurred via estrogen receptor signaling using the ER antagonist, ICI 182,780 (fulvestrant). Interestingly we found that the combination of ICI with estrogen or BPA greatly affected the expression of Set8, even when the hormone alone had no effect. This study demonstrates that the effects of estrogen and BPA on HME expression vary and that the presence of both the estrogen receptor and androgen receptor may be important for therapeutic intervention.

SET8 expression is associated with overall survival in gastric cancer.

SET8, a member of the SET domain-containing methyl-transferase, has been implicated in various biological processes. In this study, SET8 was immunostained in 100 samples of gastric cancer tissues and semi-quantified using the HSCORE method to determine the predictive value of SET8 expression levels for gastric cancer outcome. The relationship between SET8 expression and the 5-year survival rate of gastric cancer patients was assessed. High expression of SET8 was associated with a shorter survival time in gastric cancer patients, and the level of SET8 expression was found to be an independent predictor of gastric cancer outcome (relative risk = 1.939; 95% confidence interval = 1.025-3.668; P = 0.042). Analysis of SET8 levels may help in the identification of patient subgroups that are at high risk for poor disease outcomes.

Curcumin inhibits cell growth and invasion through up-regulation of miR-7 in pancreatic cancer cells

Accumulating evidence has revealed that a natural compound curcumin exerts its anti-tumor activity in pancreatic cancer. However, the underlying molecular mechanism remains elusive. Recently, miRNAs have been demonstrated to play a crucial role in tumorigenesis, suggesting that targeting miRNAs could be a promising approach for the treatment of human cancers. In this study, we explored whether curcumin regulates miR-7, leading to the inhibition of cell growth, migration and invasion in pancreatic cancer cells. We observed that curcumin suppressed cell growth, migration and invasion, and induced cell apoptosis, which is associated with increased expression of miR-7 and subsequently decreased expression of SET8, one of the miR-7 targets. These findings demonstrated that targeting miR-7 by curcumin could be a novel strategy for the treatment of pancreatic cancer.

Differential expression patterns of histone modifying enzymes in ovarian cancer in response to estradiol and bisphenol A (942.3)

Ovarian cancer is the ninth most common cancer but the fifth leading cause of cancer death for women. It has long been known that ovarian cancer growth and development is dependent upon estrogen. Conversely there is less known about how environmental estrogens influence ovarian cancer development. Estrogenic compounds have been shown to affect the epigenome, yet little is known about the specific mechanisms by which these changes occur. This work focused on the effects of both estradiol and the endocrine disruptor bisphenol A (BPA) on the expression of histone modifying enzymes (HMEs) in two different human ovarian cancer model systems, SKOV3 and OVCAR3. Specifically, the expression of Set8, a histone methyl transferase, and Sirt1, a histone deacetylase, was examined in response to physiological doses of either estrogen or BPA. In addition, the experiments were carried out using physiological conditions. There are differences in the expression pattern of these genes between the two compounds and also between the two model systems. These results imply that environmental estrogens potentially impact global gene expression by changing the expression of HMEs. Furthermore these changes are different depending on the model system analyzed demonstrating a complex effect of these compounds in human ovarian cancer.

The histone methyltransferase SET8 is required for S-phase progression

Chromatin structure and function is influenced by histone posttranslational modifications. SET8 (also known as PR-Set7 and SETD8) is a histone methyltransferase that monomethylates histonfe H4-K20. However, a function for SET8 in mammalian cell proliferation has not been determined. We show that small interfering RNA inhibition of SET8 expression leads to decreased cell proliferation and accumulation of cells in S phase. This is accompanied by DNA double-strand break (DSB) induction and recruitment of the DNA repair proteins replication protein A, Rad51, and 53BP1 to damaged regions. SET8 depletion causes DNA damage specifically during replication, which induces a Chk1-mediated S-phase checkpoint. Furthermore, we find that SET8 interacts with proliferating cell nuclear antigen through a conserved motif, and SET8 is required for DNA replication fork progression. Finally, codepletion of Rad51, an important homologous recombination repair protein, abrogates the DNA damage after SET8 depletion. Overall, we show that SET8 is essential for genomic stability in mammalian cells and that decreased expression of SET8 results in DNA damage and Chk1-dependent S-phase arrest.