Progesterone receptor membrane component-1 regulates hepcidin biosynthesis
Xiang Li, … , Donald B. Bloch, Randall T. Peterson
Xiang Li, … , Donald B. Bloch, Randall T. Peterson
Published December 14, 2015
Citation Information: J Clin Invest. 2016;126(1):389-401. https://doi.org/10.1172/JCI83831.
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Research Article Hematology

Progesterone receptor membrane component-1 regulates hepcidin biosynthesis

Abstract

Iron homeostasis is tightly regulated by the membrane iron exporter ferroportin and its regulatory peptide hormone hepcidin. The hepcidin/ferroportin axis is considered a promising therapeutic target for the treatment of diseases of iron overload or deficiency. Here, we conducted a chemical screen in zebrafish to identify small molecules that decrease ferroportin protein levels. The chemical screen led to the identification of 3 steroid molecules, epitiostanol, progesterone, and mifepristone, which decrease ferroportin levels by increasing the biosynthesis of hepcidin. These hepcidin-inducing steroids (HISs) did not activate known hepcidin-inducing pathways, including the BMP and JAK/STAT3 pathways. Progesterone receptor membrane component-1 (PGRMC1) was required for HIS-dependent increases in hepcidin biosynthesis, as PGRMC1 depletion in cultured hepatoma cells and zebrafish blocked the ability of HISs to increase hepcidin mRNA levels. Neutralizing antibodies directed against PGRMC1 attenuated the ability of HISs to induce hepcidin gene expression. Inhibiting the kinases of the SRC family, which are downstream of PGRMC1, blocked the ability of HISs to increase hepcidin mRNA levels. Furthermore, HIS treatment increased hepcidin biosynthesis in mice and humans. Together, these data indicate that PGRMC1 regulates hepcidin gene expression through an evolutionarily conserved mechanism. These studies have identified drug candidates and potential therapeutic targets for the treatment of diseases of abnormal iron metabolism.

Authors

Xiang Li, David K. Rhee, Rajeev Malhotra, Claire Mayeur, Liam A. Hurst, Emily Ager, Georgia Shelton, Yael Kramer, David McCulloh, David Keefe, Kenneth D. Bloch, Donald B. Bloch, Randall T. Peterson

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Figure 1

A small-molecule screen in transgenic zebrafish identifies compounds that promote the degradation of ferroportin.

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A small-molecule screen in transgenic zebrafish identifies compounds tha...
(A–D) Fluorescent dissection microscopy and confocal microscopy of Fpn-GFP transgenic zebrafish larvae treated with vehicle (0.1% DMSO) or 10 μM epitiostanol (dissolved in DMSO). Three-day-old transgenic embryos were soaked in E3 buffer containing either vehicle or epitiostanol for 12 hours before imaging (original magnification, ×4 [A and B]; ×40 [C and D]). High-magnification images of boxed regions in A and B are shown in C and D, respectively. (E–I) Iron staining of wild-type and ferroportin transgenic fish (ubi) treated with DMSO or epitiostanol (epi). Three-day-old embryos were treated with DMSO or 10 μM epitiostanol dissolved in DMSO for 12 hours and then fixed. Enhanced Prussian blue staining was then performed on each embryo. Arrows indicate iron deposition in the caudal hematopoietic tissue (original magnification, ×8 [E–H]). Iron staining was quantified by ImageJ based on the pixel intensity, as shown in I and described in the Methods. Results are expressed as mean ± SEM, *P < 0.001 ubi plus DMSO (n = 8), compared with wild-type plus DMSO (n = 8), wild-type plus epitiostanol (n = 14), or ubi plus epitiostanol (n = 13), ANOVA. (J) Chemical structure of some compounds that were tested in this screen. (K–P) Fluorescent microscopy (original magnification, ×4) of ferroportin transgenic fish treated with the steroid compounds listed in J. Three-day-old transgenic embryos were treated with vehicle (DMSO, n = 13), 10 μM epitiostanol dissolved in DMSO (n = 11), 10 μM mifepristone (n = 11), 10 μM progesterone (n = 9), or 10 μM dexamethasone (n = 13) for 12 hours, respectively. (P) The intensity of fluorescent staining was quantified for each treatment group and is further described in the Methods. Results are expressed as mean ± SEM, *P < 0.01, compared with the vehicle control, ANOVA.