Long non-coding RNA WAC antisense RNA 1 (lncRNA WAC-AS1) is involved in the replication of the hepatitis B virus (HBV). The purpose of this study was to determine its functions and specific mechanism. The levels of lncRNA WAC-AS1, RNA (miR)-192-5p and were examined in serum of HBV-infected patients and in HepG2.2.15 cells using quantitative reverse transcriptase polymerase chain reaction (qRT-PCR) and Western blotting. Using the database starBase, the target binding sites of lncRNA WAC-AS1 and miR-192-5p were predicted and confirmed by dual-luciferase reporter assay and RNA pull-down assay. The expression of pgRNA and HBV DNA was determined by qRT-PCR, while the levels of HBeAg and HBsAg were measured by enzyme-linked immunosorbent assay (ELISA). Using laser scanning confocal microscopy, the light chain 3 (LC3) expression was analyzed. qRT-PCR and Western blotting were used to assess the expression of beclin-1, p62, and LC3I/II. Overexpression of lncRNA WAC-AS1, upregulation of ATG7. and downregulation of miR-192-5p were observed in the serum of HBV-infected patients and the in vitro model. miR-192-5p directly targets lncRNA WAC-AS1. LncRNA WAC-AS1 was downregulated in lncRNA WAC-AS1-shRNA‒transfected cells. miR-192-5p was upregulated in lncRNA WAC-AS1-shRNA-transfected cells and downregulated in cells transfected with a miR-192-5p inhibitor. In HepG2 2.15 cells, the downregulation of lncRNA WAC-AS1 inhibited HBV replication and autophagy. In contrast, the miR-192-5p inhibitor-transfected group exhibited the opposite results, and ATG7 overexpression reversed the effects of miR-192-5p mimic or lncRNA WAC-AS1-shRNA on HBV replication and cell autophagy. Our findings indicate that lncRNA WAC-AS1 regulates HBV replication by reinforcing the autophagy induced by miR-192-5p/ATG7. Consequently, lncRNA WAC-AS1 may serve as a therapeutically-promising target in HBV patients.

中文翻译：

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长链非编码 RNA WAC 反义 RNA 1 通过增强 miR-192-5p/ATG7 诱导的自噬介导体外乙型肝炎病毒复制。

长链非编码 RNA WAC 反义 RNA 1 (lncRNA WAC-AS1) 参与乙型肝炎病毒 (HBV) 的复制。本研究的目的是确定其功能和具体机制。使用定量逆转录酶聚合酶链反应 (qRT-PCR) 和蛋白质印迹法检测 HBV 感染患者的血清和 HepG2.2.15 细胞中 lncRNA WAC-AS1、RNA (miR)-192-5p 的水平。使用数据库starBase，通过双荧光素酶报告基因分析和RNA下拉分析预测和确认lncRNA WAC-AS1和miR-192-5p的靶结合位点。qRT-PCR检测pgRNA和HBV DNA的表达，酶联免疫吸附试验（ELISA）检测HBeAg和HBsAg的水平。使用激光扫描共聚焦显微镜，分析轻链 3 (LC3) 的表达。qRT-PCR 和蛋白质印迹用于评估 beclin-1、p62 和 LC3I/II 的表达。lncRNA WAC-AS1过表达，ATG7上调。在HBV感染患者的血清和体外模型中观察到miR-192-5p的下调。miR-192-5p 直接靶向 lncRNA WAC-AS1。LncRNA WAC-AS1 在 lncRNA WAC-AS1-shRNA-转染的细胞中下调。miR-192-5p 在 lncRNA WAC-AS1-shRNA 转染的细胞中上调，在转染 miR-192-5p 抑制剂的细胞中下调。在 HepG2 2.15 细胞中，lncRNA WAC-AS1 的下调抑制了 HBV 复制和自噬。相比之下，转染 miR-192-5p 抑制剂的组表现出相反的结果，ATG7 过表达逆转了 miR-192-5p 模拟物或 lncRNA WAC-AS1-shRNA 对 HBV 复制和细胞自噬的影响。我们的研究结果表明，lncRNA WAC-AS1 通过增强 miR-192-5p/ATG7 诱导的自噬来调节 HBV 复制。因此，lncRNA WAC-AS1 可作为 HBV 患者的治疗靶点。