ATM is required for the prolactin-induced hsp90-mediated increase in cellular viability and clonogenic growth after DNA damage

Abstract

Prolactin (PRL) acts as a survival factor for breast cancer cells, but the PRL signaling pathway and the mechanism are unknown. Previously, we identified the master chaperone, heat shock protein 90 (HSP90) a, as a prolactin-Janus kinase 2 (JAK2)-signal transducer and activator of transcription 5 (STAT5) target gene involved in survival, and here we investigated the role of HSP90 in the mechanism of PRL-induced viability in response to DNA damage. The ataxia-telangiectasia mutated kinase (ATM) protein plays a critical role in the cellular response to double-strand DNA damage. We observed that PRL increased viability of breast cancer cells treated with doxorubicin or etoposide. The increase in cellular resistance is specific to the PRL receptor, because the PRL receptor antagonist, D1-9-G129R-hPRL, prevented the increase in viability. Two different HSP90 inhibitors, 17- allylamino-17-demethoxygeldanamycin and BIIB021, reduced the PRL-mediated increase in cell viability of doxorubicin-treated cells and led to a decrease in JAK2, ATM, and phosphorylatedATMprotein levels. Inhibitors of JAK2 (G6) and ATM (KU55933) abolished the PRL-mediated increase in cell viability of DNAdamaged cells, supporting the involvement of each, as well as the crosstalk of ATMwith the PRL pathway in the context ofDNAdamage. Drug synergismwas detected between theATMinhibitor (KU55933) and doxorubicin and between the HSP90 inhibitor (BIIB021) and doxorubicin. Short interfering RNA directed against ATMprevented the PRL-mediated increase in cell survival in two-dimensional cell culture, threedimensional collagen gel cultures, and clonogenic cell survival, after doxorubicin treatment. Our results indicate that ATM contributes to the PRL-JAK2-STAT5-HSP90 pathway in mediating cellular resistance to DNA-damaging agents.