Abstract
Background: Nuclear protein-1 (NUPR1, also known as p8/Com-1) is a transcription factor involved in the regulation of cellular stress responses, including serum starvation and drug stimulation. Methods: We investigated the mechanism of NUPR1 nuclear translocation involving karyopherin β1 (KPNB1), using a single-molecule binding assay and confocal microscopy. The cellular effects associated with NUPR1–KPNB1 inhibition were investigated by gene expression profiling and cell cycle analysis. Results: The single-molecule binding assay revealed that KPNB1 bound to NUPR1 with a binding affinity of 0.75 nM and that this binding was blocked by the aminothiazole ATZ-502. Following doxorubicin-only treatment, NUPR1 was translocated to the nucleus in more than 90% and NUPR1 translocation was blocked by the ATZ-502 combination treatment in MDA-MB-231 with no change in NUPR1 expression, providing strong evidence that NUPR1 nuclear translocation was directly inhibited by the ATZ-502 treatment. Inhibition of KPNB1 and NUPR1 binding was associated with a synergistic anticancer effect (up to 19.6-fold) in various cancer cell lines. NUPR1-related genes were also downregulated following the doxorubicin–ATZ-502 combination treatment. Conclusion: Our current findings clearly demonstrate that NUPR1 translocation into the nucleus requires karyopherin β1 binding. Inhibition of the KPNB1 and NUPR1 interaction may constitute a new cancer therapeutic approach that can increase the drug efficacy while reducing the side effects.
Highlights
The transcription factor nuclear protein-1 (NUPR1, aka p8/Com-1) is a member of the high-mobility group of transcriptional regulators [1], and it plays an important role in several different biological processes, including transcription, cell cycle, autophagy, and cell death [2,3,4]
MDA-MB 231 cells were treated with doxorubicin for 24, 48, and 72 h to investigate the effects of doxorubicin cytotoxicity
We established the importance of the protein–protein binding interaction between NUPR1 and karyopherin β1 in the protein nuclear translocation mechanism, and we demonstrated that ATZ-502 inhibited this interaction
Summary
The transcription factor nuclear protein-1 (NUPR1, aka p8/Com-1) is a member of the high-mobility group of transcriptional regulators [1], and it plays an important role in several different biological processes, including transcription, cell cycle, autophagy, and cell death [2,3,4]. NUPR1 is a component of the cellular stress response, playing a role in serum starvation, oxidative stress, and drug stimulation, and it is overexpressed in various cancers [5,6,7]. NUPR1 is involved in the translocation of p27 from the nucleus to cytoplasm in pancreatic cells. Nuclear protein-1 (NUPR1, known as p8/Com-1) is a transcription factor involved in the regulation of cellular stress responses, including serum starvation and drug stimulation. Methods: We investigated the mechanism of NUPR1 nuclear translocation involving karyopherin β1 (KPNB1), using a single-molecule binding assay and confocal microscopy. Inhibition of KPNB1 and NUPR1 binding was associated with a synergistic anticancer effect (up to 19.6-fold) in various cancer cell lines. Inhibition of the KPNB1 and NUPR1 interaction may constitute a new cancer therapeutic approach that can increase the drug efficacy while reducing the side effects
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