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A novel metadherinΔ7 splice variant enhances triple negative breast cancer aggressiveness by modulating mitochondrial function via NFĸB-SIRT3 axis

Oncogene volume 39, pages 2088–2102 (2020)Cite this article

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Abstract

Metadherin (MTDH) expression inversely correlates with prognosis of several cancers including mammary carcinomas. In this work, we identified a novel splice variant of MTDH with exon7 skipping (MTDHΔ7) and its levels were found significantly high in triple negative breast cancer (TNBC) cells and in patients diagnosed with TNBC. Selective overexpression of MTDHΔ7 in MDA-MB-231 and BT-549 cells enhanced proliferation, invasion, and epithelial-to-mesenchymal (EMT) transition markers in comparison to its wildtype counterpart. In contrast, knockdown of MTDHΔ7 induced antiproliferative/antiinvasive effects. Mechanistically, MTDH-NFĸB-p65 complex activated SIRT3 transcription by binding to its promoter that in turn enhanced MnSOD levels and promoted EMT in TNBC cells. Intriguingly, mitochondrial OCR through Complex-I and -IV, and glycolytic rate (ECAR) were significantly high in MDA-MB-231 cells stably expressing MTDHΔ7. While depletion of SIRT3 inhibited MTDH-Wt/Δ7-induced OCR and ECAR, knockdown of MnSOD inhibited only ECAR. In addition, MTDH-Wt/Δ7-mediated pro-proliferative/-invasive effects were greatly obviated with either siSIRT3 or siMnSOD in these cells. The functional relevance of MTDHΔ7 was further proved under in vivo conditions in an orthotopic mouse model of breast cancer. Mice bearing labeled MDA-MB-231 cells stably expressing MTDHΔ7 showed significantly more tumor growth and metastatic ability to various organs in comparison to MTDH-Wt bearing mice. Taken together, MTDHΔ7 promotes TNBC aggressiveness through enhanced mitochondrial biogenesis/function, which perhaps serves as a biomarker.

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Fig. 1: Detection of MTDHΔ7 variant in human breast cancer (BC) tissue samples and BC cell lines.
Fig. 2: MTDH-Wt and MTDHΔ7 overexpression promotes cell proliferation, invasion and migration in TNBC cells but not in normal mammary epithelial cells.
Fig. 3: MTDH-Wt/ MTDHΔ7 bind to NFĸB-p65 and translocate to nucleus and promote EMT through activation of SIRT3-MnSOD axis.
Fig. 4: MTDHΔ7 in comparison to MTDH-Wt enhances mitochondrial oxidative phosphorylation and glycolytic flux in TNBC cells: Effect of SIRT3 and MnSOD depletion.
Fig. 5: MTDH-Wt/MTDHΔ7 enhances mitochondrial oxidative phosphorylation through activation of ETC Complex-I and complex-IV: Effect of MTDH/MTDHΔ7 on mitochondrial biogenesis.
Fig. 6: MTDHΔ7 in comparison to MTDH-Wt accelerates breast tumor growth, and metastasis. Detailed animal experimental protocol is mentioned in the methods.
Fig. 7

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Acknowledgements

This work was supported by grants from the Department of Biotechnology, Department of Science and Technology, and Council of Scientific and Industrial Research, India. PKN, PNG and SM acknowledge ICMR, UGC and CSIR, for the award of a research fellowship. The authors thank Dr Madhusudhana Kuncha, Ms Sravya Panangipalli, Mr Subbarao Tulimilli, Mr Kanaka Raju Yellusani, and Mr Suresh Yerramsetty for their excellent technical support during animal experimentation, IVIS live imaging, and cell cycle analysis. We thank Dr Nishant Jain for helping in the generation of lentiviral constructs. We thank Dr Mahesh Kumar Jerald for helping in tissue mitotic index analysis and quantification. We also thank Prof. Vishnupriya Satti and Dr Sandhya Annamaneni for providing human breast cancer tissues samples. We thank Director, CSIR-IICT (Ms. No. IICT/Pubs/2019/186) for providing all the required facilities to carry out the work.

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  1. Department of Applied Biology, CSIR-Indian Institute of Chemical Technology, Uppal Road, Hyderabad, 500007, India

    Praveen Kumar Neeli, Paradesi Naidu Gollavilli, Sreevidya Mallappa & Srigiridhar Kotamraju

  2. Academy of Scientific and Innovative Research, Training and Development Complex, Chennai, 600113, India

    Praveen Kumar Neeli, Paradesi Naidu Gollavilli, Sreevidya Mallappa & Srigiridhar Kotamraju

  3. Department of Genetics, Osmania University, Hyderabad, India

    Sai Gayathri Hari

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Contributions

PKN contributed to the experimental design, data analysis, and writing of the paper. PNG was involved in the initial identification of MTDHΔ7 spliced isoform in breast cancer cells. PKN and SM contributed to immunohistopathological studies. SGH contributed for the collection of Human BC tissue samples. SK Contributed to the experimental design, provision of reagents and other material required for performing both in vitro and in vivo experiments, data analysis, and writing of the paper.

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Correspondence to Srigiridhar Kotamraju.

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Neeli, P.K., Gollavilli, P.N., Mallappa, S. et al. A novel metadherinΔ7 splice variant enhances triple negative breast cancer aggressiveness by modulating mitochondrial function via NFĸB-SIRT3 axis. Oncogene 39, 2088–2102 (2020). https://doi.org/10.1038/s41388-019-1126-6

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