Ubiquitin ligase RNF146 coordinates bone dynamics and energy metabolism
Yoshinori Matsumoto, … , Carsten Bergmann, Robert Rottapel
Yoshinori Matsumoto, … , Carsten Bergmann, Robert Rottapel
Published June 5, 2017
Citation Information: J Clin Invest. 2017;127(7):2612-2625. https://doi.org/10.1172/JCI92233.
View: Text | PDF
Research Article Bone biology Cell biology

Ubiquitin ligase RNF146 coordinates bone dynamics and energy metabolism

Abstract

Cleidocranial dysplasia (CCD) is an autosomal dominant human disorder characterized by abnormal bone development that is mainly due to defective intramembranous bone formation by osteoblasts. Here, we describe a mouse strain lacking the E3 ubiquitin ligase RNF146 that shows phenotypic similarities to CCD. Loss of RNF146 stabilized its substrate AXIN1, leading to impairment of WNT3a-induced β-catenin activation and reduced Fgf18 expression in osteoblasts. We show that FGF18 induces transcriptional coactivator with PDZ-binding motif (TAZ) expression, which is required for osteoblast proliferation and differentiation through transcriptional enhancer associate domain (TEAD) and runt-related transcription factor 2 (RUNX2) transcription factors, respectively. Finally, we demonstrate that adipogenesis is enhanced in Rnf146–/– mouse embryonic fibroblasts. Moreover, mice with loss of RNF146 within the osteoblast lineage had increased fat stores and were glucose intolerant with severe osteopenia because of defective osteoblastogenesis and subsequent impaired osteocalcin production. These findings indicate that RNF146 is required to coordinate β-catenin signaling within the osteoblast lineage during embryonic and postnatal bone development.

Authors

Yoshinori Matsumoto, Jose La Rose, Melissa Lim, Hibret A. Adissu, Napoleon Law, Xiaohong Mao, Feng Cong, Paula Mera, Gerard Karsenty, David Goltzman, Adele Changoor, Lucia Zhang, Megan Stajkowski, Marc D. Grynpas, Carsten Bergmann, Robert Rottapel

×

Figure 6

RNF146 represses adipocyte development and fat stores.

Options: View larger image (or click on image) Download as PowerPoint
RNF146 represses adipocyte development and fat stores. (A) MEFs isolated...
(A) MEFs isolated from Rnf146fl/fl (WT) and Rnf146fl/fl CMV-Cre (KO) embryos were cultured in adipogenic medium and stained with oil red O. Top panels, bright field images; bottom panels, oil red O staining. Original magnification: ×200. (B) Whole cell lysates from cells in A cultured in adipogenic medium for 3–9 days were probed with the indicated antibodies for Western blot analysis. (C and D) qPCR analysis of Pparg2 (C) or Fabp4 (D) mRNA expression in cells in A cultured in adipogenic medium for 3–9 days. n = 3. (E) Luciferase activity from a TOPflash reporter assay in cells in A cultured in serum-free medium in the presence or absence of WNT3a (40 ng/ml). n = 3. (F) Oil red O staining of MEFs in A cultured in adipogenic medium in the presence or absence of NaCl (10 mM, top panels) and LiCl (10 mM, bottom panels). Original magnification: ×200. (G) MEFs in A infected with an empty vector control (mock) or a β-catenin–expressing (S33Y) retroviral vector were cultured in adipogenic medium and stained with oil red O. Top panels, bright field images; bottom panel, oil red O staining. Original magnification: ×200. (H) H&E staining of tibiae from 12-week-old Rnf146fl/fl and Rnf146fl/fl Osx-Cre mice. Scale bar: 500 μm. (I) Quantification of trabecular bone marrow fat area in tibiae in H. n = 5–6. (J) Fat pad mass (fat pad weight over body weight) from 6-month-old Rnf146fl/fl and Rnf146fl/fl Osx-Cre mice. n = 3. P values were determined by ANOVA with Tukey-Kramer’s post-hoc test (E) or unpaired t test (C, D, I, and J). Data are presented as mean ± SEM. *P < 0.05.