CircZDHHC20 represses the proliferation, migration and invasion in trophoblast cells by miR-144/GRHL2 axis

Abstract

Background: 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. In the current study, we aimed to explore the impact and mechanisms of circRNA zinc finger DHHCtype palmitoyltransferase 20 (circZDHHC20) in PE pathogenesis. Methods: RNase R assay and reverse transcription with Oligo(dT) 18 primers were performed to confirm that circ-ZDHHC20 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-(3carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetr-azolium (MTS), wound healing and transwell assays, respectively. Western blot was used for the quantification of GRHL2 protein level. Results: Our 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. Conclusion: Our 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.

circCRAMP1L is a novel biomarker of preeclampsia risk and may play a role in preeclampsia pathogenesis via regulation of the MSP/RON axis in trophoblasts

“…Reportedly, PE has a long-term impact on cardiovascular disease development [ 26 ]; hence, early detection and intervention are particularly important. Recently, increasing evidence has suggested that circRNA is associated with PE [ 27 – 29 ].…”

Circ-PNPT1 contributes to gestational diabetes mellitus (GDM) by regulating the function of trophoblast cells through miR-889-3p/PAK1 axis

“…24 h later, the cells passing through the membrane were fixed with paraformaldehyde and stained with 0.1% crystal violet (Cat#C0121, Beyotime). Then the crystal violet-stained cells were imaged and counted (magnification 200 ×) [ 27 ].…”

“…The plates were washed with 1 mL of fresh growth medium to remove the suspended cells and then cultured in the incubator at 37 °C. Images were captured at 0 and 24 h and wound widths were quantitatively examined using a standard caliper [ 27 ].…”

“…Approximately 3 μg of total RNAs were treated without or with 3 U/μg of RNase R (Cat# RNR07250, Epicenter, Madison, WI, USA) at room temperature for 20 min, followed by purification using RNeasy MinElute Cleanup Kit (Qiagen, Tokyo, Japan). At last, the abundances of PNPT1 mRNA and circ-PNPT1 were determined using qRT-PCR assay [ 27 ].…”

Identification of mRNA-, circRNA- and lncRNA- Associated ceRNA Networks and Potential Biomarkers for Preeclampsia From Umbilical Vein Endothelial Cells

“…The higher expression of lncRNA PSG10P contributed to PE pathogenesis by regulating miR-19a-3p and IL1RAP (Wang et al, 2019). CircTRNC18 and circZDHHC20 sponge miR-762 and miR-144, respectively, affect the expression of GRHL2 and repress migration of trophoblast cells (Shen et al, 2019;Zhou et al, 2020). Due to their unique circular structure, circRNAs are resistant to ribonucleases and are more stable in body fluids or exosomes than linear RNAs.…”