| Purpose: To investigate the mechanism of SETD8 regulating the metabolic profile of breast cancer cells.Methods: Constantly SETD8-overexpressed MCF7 cell strain and constantly SETD8-knocked-down cell strain were constructed.Seahorse XF96 analyzer was employed to determine the metabolic alterations in breast cancer cells.Mass spectrometry was used to screen the proteins that are interacted with SETD8,and co-Immunoprecipitation(CoIP)and immune fluorescence assay were performed to verify the interaction between SETD8 and hypoxia-inducible-factor 1α(HIF1α).Afterwards,protein immunoblotting assay and real-time PCR were performed to determine the expression of target HIF1α genes.Dual-luciferase reporter assay was performed to determine the role of SETD8 in regulating the transcriptional activity of HIF1α and protein half-life assay was performed to detect the effect of SETD8 on HIF1α half-life.Breast cancer tissues were collected and immunohistochemistry was employed to determine the expression of SETD8 and HIF1α.At last,dual-luciferase reporter assay and real-time RCR were performed to examine the function of HIF1α in SETD8 transcription.Results: SETD8 was found to be involved in the metabolic reprogramming of tumor cells and promotes the anabolic metabolism of cancer cells.In this study,the regulation function of SETD8 in breast cancer metabolism was dependent on HIF1α.SETD8 promoted the stability of HIF1α in a posttranslational way and enhanced the transcriptional activity of HIF1α on its target genes.At the same time,we discovered that HIF1α was able to regulate the transcription of SETD8 conversely.Thus,SETD8 and HIF1α formed a positive feedback loop,corporately regulating the metabolic reprogramming and malignant biological phenotypes of breast cancer.Conclusion: In our study,we propose a novel regulator of metabolic reprogramming in tumor cells and illustrate the significant role of SETD8 in the maintenance of malignant biological behaviors of breast cancer cells. |
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