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The long non-coding RNA TP73-AS1 modulates HCC cell proliferation through miR-200a-dependent HMGB1/RAGE regulation
Shaling Li1, Yan Huang1, Yun Huang2
1Hunan Key Laboratory of Viral Hepatitis, Department of Infectious Disease, Xiangya Hospital, Central South University, Changsha, 410008, China.
Background:
P73 antisense RNA 1 T (non-protein coding), also known as TP73-AS1, is a long non-coding RNA (lncRNA) which is involved in cell proliferation and the development of tumors. However, the exact effects and molecular mechanisms of TP73-AS1 in hepatocellular carcinoma (HCC) progression are still unknown. The present study is aimed to investigate the detailed functions and the mechanism of TP73-AS1 in regulation of HCC cell proliferation.
Methods:
TP73-AS1 expression in HCC tissues and cell lines was determined using real-time PCR assays; the correlation of TP73-AS1 expression with clinicopathological features of HCC was analyzed. The functions of TP73-AS1 in regulation of HCC cell proliferation was evaluated using MTT and BrdU assays. The candidate upstream miRNAs of HMGB1 were screened using miRcode, miRWalk, miRanda and Target scan, verified using real-time PCR assays. The interaction between TP73-AS1 and miR-200a was confirmed using Luciferase report gene assays. The proten levels of HMGB1 signaling-related factors in response to co-processing TP73-AS1 knockdown and miR-200a inhibition were determined using Western blot assays and ELISA. Further, miR-200a, HMGB1 mRNA and RAGE mRNA and their correlations in HCC tissues were determined.
Results:
TP73-AS1 was upregulated in HCC tissues and cell lines. High TP73-AS1 expression was correlated with worse clinicopathological features, poorer prognosis and shorter survival. Knockdown of TP73-AS1 inhibited the HCC proliferation and the expression levels of HMGB1, RAGE and NF-κB in HCC cells. By using online tools, we screened out several candidate upstream miRNAs of HMGB1, among which miR-200a overexpression inhibited HMGB1 mRNA expression the most significantly. By using luciferase assays, we confirmed that miR-200a could directly bind to TP73-AS1 and the 3'UTR of HMGB1; TP73-AS1 competed with HMGB1 for miR-200a binding. MiR-200a inhibition could up-regulate HMGB1, RAGE, NF-κB expression as well as NF-κB regulated cytokines levels, which could be partially restored by si-TP73-AS1. In HCC tissues, miR-200a was down-regulated while HMGB1 and RAGE were up-regulated; TP73-AS1 was inversely correlated with miR-200a, while positively correlated with HMGB1 and RAGE, respectively.
Conclusion:
Our data indicated that TP73-AS1 might be an oncogenic lncRNA that promoted proliferation of HCC and could be regarded as a therapeutic target in human HCC.
Insights
TP73-AS1, a long non-coding RNA, promotes hepatocellular carcinoma (HCC) cell proliferation by interacting with miR-200a and upregulating HMGB1. Targeting TP73-AS1 offers a potential therapeutic strategy for HCC.
Area of Science:
- Oncology
- Molecular Biology
- Genetics
Background:
- Hepatocellular carcinoma (HCC) progression is influenced by long non-coding RNAs (lncRNAs), but the specific role of TP73 antisense RNA 1 (TP73-AS1) remains unclear.
- Understanding the molecular mechanisms of TP73-AS1 in HCC is crucial for developing targeted therapies.
Purpose of the Study:
- To investigate the functions of TP73-AS1 in regulating HCC cell proliferation.
- To elucidate the molecular mechanism underlying TP73-AS1's role in HCC progression.
Main Methods:
- TP73-AS1 expression analysis in HCC tissues and cell lines using real-time PCR.
- Functional assays (MTT, BrdU) to assess TP73-AS1's impact on HCC cell proliferation.
- Luciferase reporter assays to confirm interactions between TP73-AS1, miR-200a, and HMGB1.
- Western blot and ELISA to analyze protein expression levels.
Main Results:
- TP73-AS1 was significantly upregulated in HCC tissues and cell lines, correlating with poor prognosis.
- Knockdown of TP73-AS1 inhibited HCC cell proliferation and reduced HMGB1, RAGE, and NF-κB expression.
- TP73-AS1 acts as a molecular sponge for miR-200a, competing with HMGB1 and promoting HCC progression.
Conclusions:
- TP73-AS1 functions as an oncogenic lncRNA promoting HCC cell proliferation.
- The TP73-AS1/miR-200a/HMGB1 axis represents a potential therapeutic target for human HCC.