Epigenetic control of phospholipase A2 receptor expression in mammary cancer cells

Abstract

BackgroundIt has recently been proposed that the M-type phospholipase A2 receptor (PLA2R1) acts as a tumour suppressor in certain malignancies including mammary cancer. Considering that DNA methylation is an important regulator of gene transcription during carcinogenesis, in the current study we analyzed the PLA2R1 expression, PLA2R1 promoter methylation, and selected micro RNA (miRNA) levels in normal human mammary epithelial cells (HMEC) and cancer cell lines.MethodsLevels of PLA2R1 and DNA methyltransferases (DNMT) specific mRNA were determined using real-time RT-PCR. Methylation specific-high resolution melting (MS-HRM) analysis was utilized to quantify the methylation degree of selected CpG sites localized in the promoter region of the PLA2R1 gene. Expression of miRNA was tested using miScript Primer Assay system.ResultsNearly complete methylation of the analyzed PLA2R1 promoter region along with PLA2R1 gene silencing was identified in MDA-MB-453 mammary cancer cells. In MCF-7 and BT-474 mammary cancer cell lines, a higher DNA methylation degree and reduced PLA2R1 expression were found in comparison with those in normal HMEC. Synergistic effects of demethylating agent (5-aza-2′-deoxycytidine) and histone deacetylase inhibitor (trichostatin A) on PLA2R1 transcription in MDA-MB-453 cells confirmed the importance of DNA methylation and histone modification in the regulation of the PLA2R1 gene expression in mammary cells. Furthermore, significant positive correlation between the expression of DNMT1 and PLA2R1 gene methylation and negative correlation between the cellular levels of hsa-mir-141, −181b, and -181d-1 and the expression of PLA2R1 were identified in the analyzed cells. Analysis of combined z-score of miR-23b, −154 and -302d demonstrated a strong and significant positive correlation with PLA2R1 expression.ConclusionsOur data indicate that (i) PLA2R1 expression in breast cancer cells is controlled by DNA methylation and histone modifications, (ii) hypermethylation of the PLA2R1 promoter region is associated with up-regulation of DNMT1, and (iii) hsa-miR-23b, −154, and −302d, as well as hsa-miR-141, −181b, and −181d-1 are potential candidates for post-transcriptional regulation of PLA2R1 expression in mammary cancer cells.

Identification and Quantification of Heterogeneously-methylated DNA Fragments Using Epiallele-sensitive Droplet Digital Polymerase Chain Reaction (EAST-ddPCR)

“…The discovery of PLA2R1 promoter hypermethylation connected with lowered PLA2R1 expression in leukemic cells indicated a tumour-suppressive role of this receptor (19). The observed PLA2R1 promoter methylation was not restricted to leukemia, but also observed in breast cancer cell lines (20) and in clear cell renal cell carcinomas (21) and is also present in prostate cancer cell lines as shown in this study.…”

The mechanism of apoliprotein A1 down-regulated by Hepatitis B virus

“…3b ). It is well known that DNA methylation is governed by the interaction of DNA methyltransferases (DNMT) whose activities are inhibited by DNA methyltransferases inhibitor 5-aza-dC [ 19 , 20 ]. A recent study demonstrated the mRNA levels of DNMT1, DNMT3A and DNMT3B in HBV-associated tissues were significantly higher than in the non-HBV-associated tissues, which may lead to hypermethylation/inactivation of p16 [ 21 ].…”

Epigenetic Mechanisms of Tamoxifen Resistance in Luminal Breast Cancer

“…The phospholipase A2 receptor (PLA2R1) acts as a tumor suppressor in certain tumors including breast cancer. PLA2R1 has been shown to be differentially expressed in normal and mammary cancer cells, and this expression is controlled by epigenetic mechanisms such as DNA methylation and histone modification [ 48 ].…”