4β-Hydroxywithanolide E Modulates Alternative Splicing of Apoptotic Genes in Human Hepatocellular Carcinoma Huh-7
Chien-Chin Lee1, Wen-Hsin Chang2, Ya-Sian Chang1,3,4
1Epigenome Research Center, China Medical University Hospital, Taichung, Taiwan.
Abstract:
Alternative splicing is a mechanism for increasing protein diversity from a limited number of genes. Studies have demonstrated that aberrant regulation in the alternative splicing of apoptotic gene transcripts may contribute to the development of cancer. In this study, we isolated 4β-Hydroxywithanolide E (4bHWE) from the traditional herb Physalis peruviana and investigated its biological effect in cancer cells. The results demonstrated that 4bHWE modulates the alternative splicing of various apoptotic genes, including HIPK3, SMAC/DIABLO, and SURVIVIN. We also discovered that the levels of SRSF1 phospho-isoform were decreased and the levels of H3K36me3 were increased in 4bHWE treatment. Knockdown experiments revealed that the splicing site selection of SMAC/DIABLO could be mediated by changes in the level of H3K36me3 in 4bHWE-treated cells. Furthermore, we extended our study to apoptosis-associated molecules, and detected increased levels of poly ADP-ribose polymerase cleavage and the active form of CASPASE-3 in 4bHWE-induced apoptosis. In vivo experiments indicated that the treatment of tumor-bearing mice with 4bHWE resulted in a marked decrease in tumor size. This study is the first to demonstrate that 4bHWE affects alternative splicing by modulating splicing factors and histone modifications, and provides a novel view of the antitumor mechanism of 4bHWE.
Insights
4β-Hydroxywithanolide E (4bHWE) from Physalis peruviana impacts cancer cell apoptosis by altering alternative splicing of key genes. This natural compound shows promise as an antitumor agent by modulating splicing factors and histone modifications.
Area of Science:
- Molecular Biology
- Cancer Research
- Pharmacology
Background:
- Alternative splicing generates protein diversity but its dysregulation in apoptotic genes is linked to cancer development.
- Physalis peruviana is a traditional herb with potential medicinal properties.
Purpose of the Study:
- To investigate the biological effects of 4β-Hydroxywithanolide E (4bHWE) isolated from Physalis peruviana in cancer cells.
- To elucidate the molecular mechanisms underlying 4bHWE's potential antitumor activity, focusing on alternative splicing and apoptosis.
Main Methods:
- Isolation of 4bHWE from Physalis peruviana.
- Treatment of cancer cells with 4bHWE and analysis of alternative splicing patterns of apoptotic genes (HIPK3, SMAC/DIABLO, SURVIVIN).
- Assessment of splicing factor (SRSF1) phosphorylation and histone modification (H3K36me3) levels.
- Knockdown experiments to investigate the role of H3K36me3 in SMAC/DIABLO splicing.
- Analysis of apoptosis-related molecules (PARP cleavage, CASPASE-3 activation).
- In vivo studies using tumor-bearing mice to evaluate 4bHWE's efficacy.
Main Results:
- 4bHWE modulated the alternative splicing of apoptotic genes HIPK3, SMAC/DIABLO, and SURVIVIN.
- Treatment with 4bHWE decreased SRSF1 phospho-isoform levels and increased H3K36me3 levels.
- H3K36me3 levels were found to mediate SMAC/DIABLO splicing site selection in 4bHWE-treated cells.
- 4bHWE induced apoptosis, evidenced by increased PARP cleavage and active CASPASE-3.
- In vivo administration of 4bHWE significantly reduced tumor size in mice.
Conclusions:
- 4bHWE exerts antitumor effects by modulating alternative splicing through effects on splicing factors and histone modifications.
- This study provides a novel perspective on the anticancer mechanisms of 4bHWE, highlighting its impact on gene splicing and apoptosis.
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