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Updated: Dec 10, 2025
Biotin-based Pulldown Assay to Validate mRNA Targets of Cellular miRNAs
Published on: June 12, 2018
Validation of target proteins of down-regulated miR-221-5p in HeLa cells treated with Polyalthia longifolia leaf
Shanmugapriya1, Nurulhasanah Othman1, Sreenivasan Sasidharan1
1Institute for Research in Molecular Medicine (INFORMM), Universiti Sains Malaysia (USM), 11800 Gelugor, Pulau Pinang Malaysia.
Abstract:
The current study was conducted to validate the target proteins of down-regulated miR-221-5p in HeLa cells treated with P. longifolia leaf extract. The validation was done by label-free quantitative proteomics approaches, Gene Ontology (GO) and protein-protein interaction analyses after the cells transfected with miRNA mimics or miRNA inhibitor. The LC-ESI-MS/MS identified a total of 1061, 668, 564 and 940 proteins from untransfected and untreated HeLa cells, untransfected P. longifolia leaf extract-treated HeLa cells, miR-221-5p mimic-transfected P. longifolia leaf extract-treated HeLa cells and anti-miR-221-5p-transfected P. longifolia leaf extract-treated HeLa cells, respectively. The proteomic, GO and protein-protein interaction analyses showed that P. longifolia treatment regulated various protein expressions in HeLa cells, namely tropomyosin, PRKC apoptosis WT1 regulator protein (PAWR), alpha-enolase and beta-enolase, which induced apoptotic cell death after the down-regulation of miR-221-5p. Conclusively, this study showed P. longifolia leaf extract's vital contribution in regulating various protein expressions in HeLa cervical cancer cells to induce apoptotic cell death after downregulation miR-221-5p.
