Schnurri-3 regulates ERK downstream of WNT signaling in osteoblasts
Jae-Hyuck Shim, … , Laurie H. Glimcher, Dallas C. Jones
Jae-Hyuck Shim, … , Laurie H. Glimcher, Dallas C. Jones
Published August 15, 2013
Citation Information: J Clin Invest. 2013;123(9):4010-4022. https://doi.org/10.1172/JCI69443.
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Research Article Bone biology

Schnurri-3 regulates ERK downstream of WNT signaling in osteoblasts

Abstract

Mice deficient in Schnurri-3 (SHN3; also known as HIVEP3) display increased bone formation, but harnessing this observation for therapeutic benefit requires an improved understanding of how SHN3 functions in osteoblasts. Here we identified SHN3 as a dampener of ERK activity that functions in part downstream of WNT signaling in osteoblasts. A D-domain motif within SHN3 mediated the interaction with and inhibition of ERK activity and osteoblast differentiation, and knockin of a mutation in Shn3 that abolishes this interaction resulted in aberrant ERK activation and consequent osteoblast hyperactivity in vivo. Additionally, in vivo genetic interaction studies demonstrated that crossing to Lrp5–/– mice partially rescued the osteosclerotic phenotype of Shn3–/– mice; mechanistically, this corresponded to the ability of SHN3 to inhibit ERK-mediated suppression of GSK3β. Inducible knockdown of Shn3 in adult mice resulted in a high–bone mass phenotype, providing evidence that transient blockade of these pathways in adults holds promise as a therapy for osteoporosis.

Authors

Jae-Hyuck Shim, Matthew B. Greenblatt, Weiguo Zou, Zhiwei Huang, Marc N. Wein, Nicholas Brady, Dorothy Hu, Jean Charron, Heather R. Brodkin, Gregory A. Petsko, Dennis Zaller, Bo Zhai, Steven Gygi, Laurie H. Glimcher, Dallas C. Jones

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Figure 5

SHN3 regulates GSK3β activity and β-catenin level in osteoblasts.

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SHN3 regulates GSK3β activity and β-catenin level in osteoblasts. (A) Pr...
(A) Primary WT COBs were stimulated with murine WNT3a (200 ng/ml) for the indicated time points after 12 hour serum starvation. Cells were lysed and immunoblotted with antibodies specific to phospho-ERK and GAPDH. (B) HEK293 cells were transfected with a xWNT8/Fz5 fusion protein along with LRP5, lysed, and immunoprecipitated by anti-ERK1/2 antibody and protein A–agarose. The immunoprecipitates were incubated with GST-ELK1 and either His-SHN3-WT or His-SHN3-KA, and ERK kinase activity was analyzed by in vitro kinase assay. Inputs indicate loading control of recombinant His-SHN3. (C and E) Primary Shn3+/+ and Shn3–/– (C) or Shn3KI/KI (E) COBs were cultured in differentiation medium, lysed, and immunoblotted with the indicated antibodies. (D) Primary Shn3–/– COBs were infected with lentivirus expressing vector or SHN3 and cultured in differentiation medium containing puromycin for 6 days. Cells were lysed and immunoblotted with the indicated antibodies. (F) SHN3 regulation of ERK activity in the context of WNT/β-catenin signaling.