Micro‐Ribonucleic Acid 494 regulation of protein S expression

Abstract

Summary. Background: Acquired protein S (PS) deficiency is highly associated with elevated circulating estrogen levels resulting from pregnancy, oral contraceptives, and estrogen replacement therapy; however, the mechanism of estrogen-mediated acquired PS deficiency remains poorly understood. Increasing evidence indicates that estrogen receptor signaling can indirectly modulate the expression of target genes at the post-transcriptional level by modulating the expression of microRNAs (miRNAs), and miRNAs have also been demonstrated to be involved in the regulation of hemostasis. Objectives: To investigate the mechanism of estrogen-mediated downregulation of PROS1 expression by the microRNA miR-494. Methods: Computational analyses of the PROS1 3′-untranslated region (UTR) were performed to identify putative miRNA-binding sites, and direct targeting of the PROS1 3′-UTR by miR-494 was determined with dual luciferase reporter assays in HuH-7 cells. Reporter vectors containing the PROS1 3′-UTR sequence with deleted miR-494-binding sites were also analyzed with luciferase reporter assays. The effects of estrogen on miR-494 and PROS1 mRNA levels in HuH-7 cells were determined by quantitative real-time PCR, and estrogen-mediated changes to secreted PS levels in culture supernatant of HuH-7 cells were measured with an ELISA. Results: The PROS1 3′-UTR sequence contains three putative miR-494-binding sites. miR-494 directly targets PROS1, and miR-494 levels are upregulated following estrogen treatment in HuH-7 liver cells in association with downregulated PROS1 mRNA and PS levels. Conclusions: The results from this study provide the first evidence for miRNA downregulation of PROS1 by miR-494, and suggest that miR-494 is involved in the mechanism of estrogen-mediated downregulation of PS expression.

The role of microRNA‐27a/b and microRNA‐494 in estrogen‐mediated downregulation of tissue factor pathway inhibitor α

“…miRNA expression has been detected in anucleate platelets [24,25], exosomes [26], and microparticles [27] possessing procoagulant properties. Moreover, recent reports have demonstrated that miRNAs directly regulate hemostatic proteins, such as protein S, antithrombin, tissue factor, fibrinogen, PAI-1, and FXI [21,[28][29][30][31][32]. Collectively, these studies suggest that miRNAs play a role in haemostasis and could contribute to the pathogenesis of thrombotic disorders.…”

“…In our study, 1-10 nmol L -1 EE2 was found sufficient to induce miR-27a and miR-27b expression levels in MCF7 cells, an effect that was evidently exerted through ERa as blocking the receptor with the ERa antagonist fulvestrant, or indeed transiently knock down of the receptor by siRNA, led to loss of the EE2-mediated induction of miR-27a/b and miR-494. In a recent study by Tay et al [28], miR-494 was reported to be induced by 1-10 nmol Fig. 6.…”

“…In a recent study by Tay et al . , miR‐494 was reported to be induced by 1–10 nmol L –1 E2 treatment in HuH‐7 cells that occurred in association with a dose‐dependent decrease in PROS1 mRNA and protein S protein levels. Protein S is a known cofactor for TFPIα and potentiates its inhibitory effect .…”

Identification of reference miRNAs in plasma useful for the study of oestrogen-responsive miRNAs associated with acquired Protein S deficiency in pregnancy

“…We demonstrated in HuH-7 liver carcinoma cells that oestrogen treatment resulted in the induction of miR-494 expression in association with downregulated levels of the anticoagulant, Protein S, indicating a mechanism for miRNA-mediated regulation of acquired Protein S deficiency, with a potential role for increased miR-494 levels in pregnant women as an indicator for high thrombotic risk [17]. It is likely that increased circulating oestrogen levels during pregnancy alters the expression of a suite of miRNAs in oestrogen-sensitive tissues, with changes in specific miRNAs possibly detectable in circulating plasma.…”

Regulation of Coagulation Factor XI Expression by MicroRNAs in the Human Liver

“…Coagulation factors like fibrinogen [22] or tissue factor have been described as interacting with miRNAs, which may have an impact on the thrombotic etiology associated with pathologies such as antiphospholipid syndrome or systemic lupus erythematosus [23]. Recently, PAI-1 [24] and protein S [25] have also been shown to be directly regulated by miRNAs. In the current study, we investigated the potential relevance of miRNAs aiming to discover new elements that may modulate FXI in the liver.…”