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IGFBP-3 interacts with NONO and SFPQ in PARP-dependent DNA damage repair in triple-negative breast cancer

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Abstract

Women with triple-negative breast cancer (TNBC) are generally treated by chemotherapy but their responsiveness may be blunted by DNA double-strand break (DSB) repair. We previously reported that IGFBP-3 forms nuclear complexes with EGFR and DNA-dependent protein kinase (DNA-PKcs) to modulate DSB repair by non-homologous end-joining (NHEJ) in TNBC cells. To discover IGFBP-3 binding partners involved in chemoresistance through stimulation of DSB repair, we analyzed the IGFBP-3 interactome by LC–MS/MS and confirmed interactions by coimmunoprecipitation and proximity ligation assay. Functional effects were demonstrated by DNA end-joining in vitro and measurement of γH2AX foci. In response to 20 µM etoposide, the DNA/RNA-binding protein, non-POU domain-containing octamer-binding protein (NONO) and its dimerization partner splicing factor, proline/glutamine-rich (SFPQ) formed complexes with IGFBP-3, demonstrated in basal-like TNBC cell lines HCC1806 and MDA-MB-468. NONO binding to IGFBP-3 was also shown in a cell-free biochemical assay. IGFBP-3 complexes with NONO and SFPQ were blocked by inhibiting EGFR with gefitinib or DNA-PKcs with NU7026, and by the PARP inhibitors veliparib and olaparib, which also reduced DNA end-joining activity and delayed the resolution of the γH2AX signal (i.e. inhibited DNA DSB repair). Downregulation of the long noncoding RNA in NHEJ pathway 1 (LINP1) by siRNA also blocked IGFBP-3 interaction with NONO–SFPQ. These findings suggest a PARP-dependent role for NONO and SFPQ in IGFBP-3-dependent DSB repair and the involvement of LINP1 in the complex formation. We propose that targeting of the DNA repair function of IGFBP-3 may enhance chemosensitivity in basal-like TNBC, thus improving patient outcomes.

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Acknowledgements

This work was supported by Grants DP140100137 from the Australian Research Council and IN-17-040 from the National Breast Cancer Foundation, Australia, to RCB.

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  1. Mike Z. Lin

    Present address: Orange Family Medical Centre, 95 Peisley Street, Orange, NSW, 2800, Australia

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  1. Kolling Institute, Royal North Shore Hospital, The University of Sydney, St. Leonards, NSW, 2065, Australia

    Hasanthi C. de Silva, Mike Z. Lin, Leo Phillips, Janet L. Martin & Robert C. Baxter

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Contributions

The study was designed by RCB. Data were collected and analysed by HCdS, MZL, LP, JLM and RCB. The manuscript was drafted by RCB. All authors revised the article critically for intellectual content and gave their approval of this version to be published.

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Correspondence to Robert C. Baxter.

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de Silva, H.C., Lin, M.Z., Phillips, L. et al. IGFBP-3 interacts with NONO and SFPQ in PARP-dependent DNA damage repair in triple-negative breast cancer. Cell. Mol. Life Sci. 76, 2015–2030 (2019). https://doi.org/10.1007/s00018-019-03033-4

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