{"title":"长非编码 RNA WAC 反义 RNA 1 通过加强 miR-192-5p/ATG7 诱导的自噬作用,在体外介导乙型肝炎病毒的复制。","authors":"Minkai Cao, Deping Yuan, Hongxiu Jiang, Guanlun Zhou, Chao Chen, Guorong Han","doi":"10.4081/ejh.2022.3438","DOIUrl":null,"url":null,"abstract":"<p><p>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.</p>","PeriodicalId":50487,"journal":{"name":"European Journal of Histochemistry","volume":null,"pages":null},"PeriodicalIF":2.1000,"publicationDate":"2022-09-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/44/92/ejh-66-3-3438.PMC9471915.pdf","citationCount":"1","resultStr":"{\"title\":\"Long non-coding RNA WAC antisense RNA 1 mediates hepatitis B virus replication <em>in vitro</em> by reinforcing miR-192-5p/ATG7-induced autophagy.\",\"authors\":\"Minkai Cao, Deping Yuan, Hongxiu Jiang, Guanlun Zhou, Chao Chen, Guorong Han\",\"doi\":\"10.4081/ejh.2022.3438\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><p>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.</p>\",\"PeriodicalId\":50487,\"journal\":{\"name\":\"European Journal of Histochemistry\",\"volume\":null,\"pages\":null},\"PeriodicalIF\":2.1000,\"publicationDate\":\"2022-09-02\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://ftp.ncbi.nlm.nih.gov/pub/pmc/oa_pdf/44/92/ejh-66-3-3438.PMC9471915.pdf\",\"citationCount\":\"1\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"European Journal of Histochemistry\",\"FirstCategoryId\":\"99\",\"ListUrlMain\":\"https://doi.org/10.4081/ejh.2022.3438\",\"RegionNum\":4,\"RegionCategory\":\"生物学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q4\",\"JCRName\":\"CELL BIOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"European Journal of Histochemistry","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.4081/ejh.2022.3438","RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q4","JCRName":"CELL BIOLOGY","Score":null,"Total":0}
Long non-coding RNA WAC antisense RNA 1 mediates hepatitis B virus replication in vitro by reinforcing miR-192-5p/ATG7-induced autophagy.
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.
期刊介绍:
The Journal publishes original papers concerning investigations by histochemical and immunohistochemical methods, and performed with the aid of light, super-resolution and electron microscopy, cytometry and imaging techniques. Coverage extends to:
functional cell and tissue biology in animals and plants;
cell differentiation and death;
cell-cell interaction and molecular trafficking;
biology of cell development and senescence;
nerve and muscle cell biology;
cellular basis of diseases.
The histochemical approach is nowadays essentially aimed at locating molecules in the very place where they exert their biological roles, and at describing dynamically specific chemical activities in living cells. Basic research on cell functional organization is essential for understanding the mechanisms underlying major biological processes such as differentiation, the control of tissue homeostasis, and the regulation of normal and tumor cell growth. Even more than in the past, the European Journal of Histochemistry, as a journal of functional cytology, represents the venue where cell scientists may present and discuss their original results, technical improvements and theories.