Primary and acquired drug resistance is one of the main obstacles encountered in high-grade serous ovarian cancer (HGSC) chemotherapy. Cisplatin induces DNA damage through cross-linking and long integrated non-coding RNAs (lincRNAs) play an important role in chemical induced DNA-damage response, which suggests that lincRNAs may be also associated with cisplatin resistance. However, the mechanism of long integrated non-coding RNAs (lincRNAs) acting on cisplatin resistance is not well understood. Here, we showed that expression of lin-RECK-3, H19, LUCAT1, LINC00961, and linc-CARS2-2 was enhanced in cisplatin-resistant A2780-DR cells, while transcriptome sequencing showed decreased Linc-TNFRSF19-1 and LINC00515 expression. Additionally, we verified that different H19 expression levels in HGSC tissues showed strong correlation with cancer recurrence. H19 knockdown in A2780-DR cells resulted in recovery of cisplatin sensitivity in vitro and in vivo. Quantitative proteomics analysis indicated that six NRF2-targeted proteins, including NQO1, GSR, G6PD, GCLC, GCLM and GSTP1 involved in the glutathione metabolism pathway, were reduced in H19-knockdown cells. Furthermore, H19-knockdown cells were markedly more sensitive to hydrogen-peroxide treatment and exhibited lower glutathione levels. Our results reveal a previously unknown link between H19 and glutathione metabolism in the regulation of cancer-drug resistance.
Lysine acetylation and succinylation are major types of protein acylation that are important in many cellular processes including gene transcription, cellular metabolism, DNA damage response. Malfunctions in these post-translational modifications are associated with genome instability and disease in higher organisms. In this study, we used high-resolution nano liquid chromatography-tandem mass spectrometry combined with affinity purification to quantify the dynamic changes of protein acetylation and succinylation in response to ultraviolet (UV)-induced cell stress. A total of 3345 acetylation sites in 1440 proteins and 567 succinylation sites in 246 proteins were identified, many of which have not been reported previously. Bioinformatics analysis revealed that these proteins are involved in many important biological processes, including cell signalling transduction, protein localization and cell metabolism. Crosstalk analysis between these two modifications indicated that modification switches might regulate protein function in response to UV-induced DNA damage. We further illustrated that FEN1 acetylation at different sites could lead to different cellular phenotypes, suggesting the multiple function involvement of FEN1 acetylation under DNA damage stress. These systematic analyses provided valuable resources and new insight into the potential role of lysine acetylation and succinylation under physiological and pathological conditions.
Ovarian cancer is one of the most lethal types of gynecological cancer. Drug resistance is a major underlying cause of treatment failure, which has lead to continued poor mortality and morbidity rates in patients. In the present study, highly sensitive transcriptome sequencing was performed to systematically identify differentially expressed mRNAs in cisplatin-sensitive (A2780) and -resistant (A2780-DR) cells. Calcium/calmodulin dependent protein kinase IIδ (CAMK2D) and SWI/SNF related matrix associated actin dependent regulator of chromatin subfamily A member 2 (SMARCA2) were identified as exhibiting increased expression in cisplatin-resistant cells. Overexpression of either SMARCA2 or CAMK2D led to a significant increase in the survival rates of A2780 and SKVO3 cells following cisplatin treatment. To further verify the contribution of these two genes in the development of drug resistance, the RNA levels in tissues with different recurrence-free survival (RFS) rates were compared. An increased mRNA level of CAMK2D was detected in samples with shorter RFS rates. An apoptosis assay revealed that overexpression of SMARCA2 or CAMK2D increased the resistance of ovarian cancer cells to cisplatin, as indicated by the decreased apoptotic cell populations. The levels of these two genes also affected the cell cycle and apoptosis-associated protein expression. Quantitative proteomic analyses revealed that overexpression of SMARCA2 or CAMK2D influences multiple metabolism and cancer-associated signaling pathways, which are critical for responses to cisplatin treatment and drug resistance development.
Scientific Reports 6: Article number: 30212; published online: 25 July 2016; updated: 26 May 2017. In the original version of this Article, Supplementary Datasets 1, 2, 3, 4 and 5 were omitted. The correct Supplementary Datasets now accompany the Article.
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