15-Deoxy-Δ^12,14-prostaglandin J₂ stabilizes hypoxia inducible factor-1α through induction of heme oxygenase-1 and direct modification ofprolyl-4-hydroxylase 2

Abstract

15-Deoxy-Δ^12,14-prostaglandin J₂ (15d-PGJ₂), a representative J-series cyclopentenone prostaglandin, has biphasic roles in cell proliferation and apoptosis. Hypoxia inducible factor-1 (HIF-1) regulates expression of various genes involved in tumor growth and angiogenesis. In the present study, treatment of human breast cancer (MCF-7) cells with 15d-PGJ₂ resulted in the accumulation of the α-subunit of HIF-1. Pretreatment with zinc protoporphyrin IX, a pharmacological inhibitor of heme oxygenase-1 (HO-1), as well as siRNA knockdown of HO-1 gene in MCF-7 cells attenuated 15d-PGJ₂-mediated HIF-1α accumulation. 15d-PGJ₂ treatment increased intracellular production of reactive oxygen species (ROS), which was mediated by HO-1 induction. Preincubation of MCF-7 cells with trolox, a water-soluble form of vitamin E, attenuated 15d-PGJ₂-induced HIF-1α expression although HO-1 expression was unchanged. This finding suggests that ROS accumulated as a consequence of HO-1 up-regulation can enhance HIF-1α expression in MCF-7 cells treated with 15d-PGJ₂. Alternatively, 15d-PGJ₂ was found to covalently bind to HIF-1α prolyl-4-hydroxylase 2 (PHD2) in MCF-7 cells, which hampers the proline hydroxylation of HIF-1α, thereby disrupting ubiquitin-dependent proteasomal degradation of this transcription factor. Pretreatment with thiol reducing agents blunted 15d-PGJ₂-induced HIF-1α stabilization, indicative of a cysteine residue as a direct target of 15d-PGJ₂. Molecular docking analysis suggests that 15d-PGJ₂ preferentially binds to PHD2 in the vicinity of the Cys²⁰¹ residue based on binding energies and carbon–sulfur distances. In summary, 15d-PGJ₂ stabilizes HIF-1α in MCF-7 cells through HO-1 induction with subsequent ROS generation and also through direct modification of PHD2.

Keywords:

• 15-Deoxy-Δ^12,14-prostaglandin J₂
• hypoxia inducible factor-1α
• heme oxygenase-1
• prolyl-4-hydroxylase 2
• MCF-7 cells
• cyclopentenone prostaglandin

Acknowledgements

The authors acknowledge the supply of the biotinylated 15d-PGJ₂, by Prof. Y.-G. Suh of Seoul National University.

Disclosure statement

The authors report no conflicts of interest. The authors alone are responsible for the content and writing of this article.

Funding

This work was supported by the Global Core Research Center (GCRC) grant [No. 2011-003-0001] from the National Research Foundation, Ministry of Education, Science and Technology, Republic of Korea.