Abstract
BackgroundPreeclampsia (PE) is a prevalent pregnancy disorder that has been one of the leading causes of maternal and perinatal mortality worldwide. Circular RNAs (circRNAs) have recently considered as important regulators in PE pathogenesis. In the current study, we aimed to explore the impact and mechanisms of circRNA zinc finger DHHC-type palmitoyltransferase 20 (circZDHHC20) in PE pathogenesis.MethodsRNase R assay and reverse transcription with Oligo(dT)18 primers were performed to confirm that circZDHHC20 was indeed circular transcript. The expression of circZDHHC20, grainyhead-like 2 (GRHL2) and miR-144 were assessed by quantitative real-time polymerase chain reaction (qRT-PCR). Subcellular localization assay was used to determine whether circZDHHC20 was predominantly present in the cytoplasm. The target correlations between miR-144 and circZDHHC20 or GRHL2 were confirmed using dual-luciferase reporter and RNA immunoprecipitation (RIP) assays. Cell proliferation, migration, and invasion were detected by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetr-azolium (MTS), wound healing and transwell assays, respectively. Western blot was used for the quantification of GRHL2 protein level.ResultsOur data indicated that circZDHHC20 was up-regulated and miR-144 was down-regulated in PE placenta. CircZDHHC20 sequestered miR-144 by acting as a miR-144 sponge. CircZDHHC20 overexpression repressed trophoblast cell proliferation, migration, and invasion, while its knockdown exerted opposite effects. Moreover, miR-144 mediated the regulation of circZDHHC20 on trophoblast cell behaviors. GRHL2 was directly targeted and inhibited by miR-144. MiR-144 exerted regulatory effects on trophoblast cell proliferation, migration and invasion by GRHL2. Furthermore, circZDHHC20 modulated GRHL2 expression through sponging miR-144.ConclusionOur study suggested that a high level of circZDHHC20 inhibited the proliferation, migration, and invasion in trophoblast cells at least partially through sponging miR-144 and up-regulating GRHL2, providing a novel mechanism of PE pathogenesis.
Highlights
Preeclampsia (PE) is a prevalent pregnancy disorder that has been one of the leading causes of maternal and perinatal mortality worldwide
To confirm that circZDHHC20 was circular transcript, RNase R assay was performed. These results revealed that linear transcript was significantly digested by RNase R and circZDHHC20 was resistant to RNase R digestion (Fig. 1b)
The data of subcellular localization assay showed that circZDHHC20 was highly enriched in the cytoplasm fraction in HTR-8/SVneo cells (Fig. 1d). qRTPCR results demonstrated that miR-144 expression was prominently reduced in placental tissues from PE patients compared to those of healthy volunteers (Fig. 1e)
Summary
Preeclampsia (PE) is a prevalent pregnancy disorder that has been one of the leading causes of maternal and perinatal mortality worldwide. Circular RNAs (circRNAs) have recently considered as important regulators in PE pathogenesis. We aimed to explore the impact and mechanisms of circRNA zinc finger DHHCtype palmitoyltransferase 20 (circZDHHC20) in PE pathogenesis. Migration, and invasion of trophoblast cells are fundamental in maintaining the function of human placenta [3]. Zhou et al [8] reported that circRNA pregnancy-associated plasma protein A (circPAPPA) was down-regulated in PE placenta, and its deficiency hampered trophoblast cell proliferation and invasion. Recent research demonstrated that circRNA zinc finger DHHC-type palmitoyltransferase 20 (circZDHHC20, hsa_circ_0006732) was up-regulated in maternal blood cells of PE patients, eliciting its potential involvement in PE pathogenesis [10]. The impact and molecular mechanisms of circZDHHC20 in PE pathogenesis remain indistinct
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