Steap4 Plays a Critical Role in Osteoclastogenesis in Vitro by Regulating Cellular Iron/Reactive Oxygen Species (ROS) Levels and cAMP Response Element-binding Protein (CREB) Activation

Jian Zhou,Shiqiao Ye,Toshifumi Fujiwara,Stavros C Manolagas,Haibo Zhao

doi:10.1074/jbc.m113.478750

Jian Zhou, Shiqiao Ye + Show 3 more

Open Access

https://doi.org/10.1074/jbc.m113.478750

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Abstract

Iron is essential for osteoclast differentiation, and iron overload in a variety of hematologic diseases is associated with excessive bone resorption. Iron uptake by osteoclast precursors via the transferrin cycle increases mitochondrial biogenesis, reactive oxygen species production, and activation of cAMP response element-binding protein, a critical transcription factor downstream of receptor activator of NF-κB-ligand-induced calcium signaling. These changes are required for the differentiation of osteoclast precursors to mature bone-resorbing osteoclasts. However, the molecular mechanisms regulating cellular iron metabolism in osteoclasts remain largely unknown. In this report, we provide evidence that Steap4, a member of the six-transmembrane epithelial antigen of prostate (Steap) family proteins, is an endosomal ferrireductase with a critical role in cellular iron utilization in osteoclasts. Specifically, we show that Steap4 is the only Steap family protein that is up-regulated during osteoclast differentiation. Knocking down Steap4 expression in vitro by lentivirus-mediated short hairpin RNAs inhibits osteoclast formation and decreases cellular ferrous iron, reactive oxygen species, and the activation of cAMP response element-binding protein. These results demonstrate that Steap4 is a critical enzyme for cellular iron uptake and utilization in osteoclasts and, thus, indispensable for osteoclast development and function.

Highlights

Iron uptake through the transferrin-dependent pathway is essential for osteoclast differentiation
Steap4 mRNA Expression Increases during Osteoclast Differentiation—In search of molecules regulating osteoclast iron metabolism, we performed a real-time PCR experiment to examine the expression of sixtransmembrane epithelial antigen of prostate (Steap) family members during oste
Steap4 mRNA was undetected in Bone marrow macrophages (BMMs), its expression was up-regulated during osteoclast differentiation, reaching its highest level in mature cells

Summary

Background

Iron uptake through the transferrin-dependent pathway is essential for osteoclast differentiation. Iron uptake by osteoclast precursors via the transferrin cycle increases mitochondrial biogenesis, reactive oxygen species production, and activation of cAMP response element-binding protein, a critical transcription factor downstream of receptor activator of NF-␬B-ligand-induced calcium signaling. These changes are required for the differentiation of osteoclast precursors to mature bone-resorbing osteoclasts. Knocking down Steap expression in vitro by lentivirus-mediated short hairpin RNAs inhibits osteoclast formation and decreases cellular ferrous iron, reactive oxygen species, and the activation of cAMP response elementbinding protein These results demonstrate that Steap is a critical enzyme for cellular iron uptake and utilization in osteoclasts and, indispensable for osteoclast development and function. PGC-1␤ acts to stimulate mitochondrion biogenesis and the generation of reactive oxygen species (ROS), which promote osteoclastogenesis by increasing the activity of CREB and PGC-1␤ in a positive feedback manner [8, 9]

The abbreviations used are

EXPERIMENTAL PROCEDURES

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DISCUSSION