A nonclassical vitamin D receptor pathway suppresses renal fibrosis
Ichiaki Ito, … , Kazuo Nagasawa, Junn Yanagisawa
Ichiaki Ito, … , Kazuo Nagasawa, Junn Yanagisawa
Published October 25, 2013
Citation Information: J Clin Invest. 2013;123(11):4579-4594. https://doi.org/10.1172/JCI67804.
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Research Article

A nonclassical vitamin D receptor pathway suppresses renal fibrosis

Abstract

The TGF-β superfamily comprises pleiotropic cytokines that regulate SMAD and non-SMAD signaling. TGF-β–SMAD signal transduction is known to be involved in tissue fibrosis, including renal fibrosis. Here, we found that 1,25-dihydroxyvitamin D3–bound [1,25(OH)2D3-bound] vitamin D receptor (VDR) specifically inhibits TGF-β–SMAD signal transduction through direct interaction with SMAD3. In mouse models of tissue fibrosis, 1,25(OH)2D3 treatment prevented renal fibrosis through the suppression of TGF-β–SMAD signal transduction. Based on the structure of the VDR-ligand complex, we generated 2 synthetic ligands. These ligands selectively inhibited TGF-β–SMAD signal transduction without activating VDR-mediated transcription and significantly attenuated renal fibrosis in mice. These results indicate that 1,25(OH)2D3-dependent suppression of TGF-β–SMAD signal transduction is independent of VDR-mediated transcriptional activity. In addition, these ligands did not cause hypercalcemia resulting from stimulation of the transcriptional activity of the VDR. Thus, our study provides a new strategy for generating chemical compounds that specifically inhibit TGF-β–SMAD signal transduction. Since TGF-β–SMAD signal transduction is reportedly involved in several disorders, our results will aid in the development of new drugs that do not cause detectable adverse effects, such as hypercalcemia.

Authors

Ichiaki Ito, Tsuyoshi Waku, Masato Aoki, Rumi Abe, Yu Nagai, Tatsuya Watanabe, Yuka Nakajima, Ichiro Ohkido, Keitaro Yokoyama, Hiroyuki Miyachi, Toshiyuki Shimizu, Akiko Murayama, Hiroyuki Kishimoto, Kazuo Nagasawa, Junn Yanagisawa

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Figure 4

H12 in VDR is required for the suppression of TGF-β–SMAD signal transduction.

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H12 in VDR is required for the suppression of TGF-β–SMAD signal transduc...
(A) The VDR-LBD and ligand-binding pocket are shown in the crystal structure of VDR complexed with 1,25(OH)2D3 (PDB: 1DB1). An enlarged view of the ligand-binding pocket is also shown. H12 is shown in magenta; 1,25(OH)2D3 is shown in orange. (B) H12 is necessary for transactivation and suppression of TGF-β signal transduction. VDR truncations are as follows: full, 1–427 aa; ΔH12, 1–403 aa; CDEF, 20–427 aa; CDE, 20–403 aa; DEF (LBD), 110–427 aa; DE (LBD-ΔH12), 110–403 aa. HEK293 cells were transfected with plasmids encoding ALK5 TD and the indicated VDR mutants and the reporter plasmids VDRE-Luc or CAGA-Luc. After culturing transfected cells with or without 1,25(OH)2D3, cell extracts were analyzed using luciferase assay. (C) H12 is necessary for SMAD3 binding. Purified recombinant His-tagged SMAD3-MH1 was incubated with GST-VDR-LBD or GST-VDR-LBD-ΔH12 in the presence or absence of 1,25(OH)2D3, after which the mixtures were examined by in vitro pulldown assay. *P < 0.05; **P < 0.01; ***P < 0.001.