Validation of target proteins of down-regulated miR-221-5p in HeLa cells treated with Polyalthia longifolia leaf extract
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ORIGINAL ARTICLE
Validation of target proteins of down‑regulated miR‑221‑5p in HeLa cells treated with Polyalthia longifolia leaf extract using label‑free quantitative proteomics approaches Shanmugapriya1 · Nurulhasanah Othman1 · Sreenivasan Sasidharan1 Received: 28 April 2020 / Accepted: 12 August 2020 © King Abdulaziz City for Science and Technology 2020
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-miR221-5p-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. Keywords miRNA · Apoptosis · HeLa cells, target protein · P. longifolia · Cancer · Natural products
Introduction The major function of miRNAs in regulating various cellular processes is mainly through partially complementary binding to their mRNA targets. The partial pairing allows a single miRNA to target some multiple genes, which leads to the regulation of gene expression at the post-transcriptional level and subsequent protein translational repression (Catalanotto et al. 2016). This substantiates the major role of miRNA in diverse biological processes such as cell death (Conti et al. 2020) and cell proliferation (Yu et al. 2018). Interestingly, various plants rich in anti-cancer bioactive compounds have been shown to regulate mammalian miRNA levels (Xie et al. Electronic supplementary material The online version of this article (https://doi.org/10.1007/s13205-020-02396-x) contains supplementary material, which is available to authorized users. * Sreenivasan Sasidharan [email protected] 1
Institute for Research in Molecular Medicine (INFORMM), Universiti Sains Malaysia (USM), 11800 Gelugor, Pulau Pinang, Malaysia
2016; Gezici and Sekeroglu 2017). Polyalthia longifolia var. Angustifolia Thw. (Annonaceae) is one of the most important indigenous medicinal plants which exert various pharmacological activities. The induction of apoptosis by P. longifolia treatment has been revealed to cause cell cycle arrest at the sub-G0/G1, G0/G1
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