IGFBP-3 interacts with NONO and SFPQ in PARP-dependent DNA damage repair in triple-negative breast cancer.
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.
['de Silva|Hasanthi C|HC|', 'Lin|Mike Z|MZ|http://orcid.org/0000-0003-1868-2495', 'Phillips|Leo|L|http://orcid.org/0000-0001-8544-8899', 'Martin|Janet L|JL|http://orcid.org/0000-0002-8039-4390', 'Baxter|Robert C|RC|http://orcid.org/0000-0001-5061-2142']
D001562:Benzimidazoles D045744:Cell Line, Tumor D059766:DNA End-Joining Repair D004268:DNA-Binding Proteins D005260:Female D015972:Gene Expression Regulation, Neoplastic D006801:Humans D018972:Insulin-Like Growth Factor Binding Protein 3 D034521:Nuclear Matrix-Associated Proteins D050810:Octamer Transcription Factors D000072670:PTB-Associated Splicing Factor D000067856:Poly(ADP-ribose) Polymerase Inhibitors D011065:Poly(ADP-ribose) Polymerases D011485:Protein Binding D034622:RNA Interference D062085:RNA, Long Noncoding D016601:RNA-Binding Proteins D064726:Triple Negative Breast Neoplasms
TNBC PARP inhibitors IGF binding protein P54NRB PSF lncRNA