High Bone Turnover in Mice Carrying a Pathogenic Notch2 Mutation Causing Hajdu-Cheney Syndrome

Abstract

Hajdu-Cheney syndrome (HCS) is a rare autosomal-dominant disorder primarily characterized by acro-osteolysis and early-onset osteoporosis. Genetically, HCS is caused by nonsense or deletion mutations within exon 34 of the NOTCH2 gene, resulting in premature translational termination and production of C-terminally truncated NOTCH2 proteins that are predicted to activate NOTCH2-dependent signaling. To understand the role of Notch2 in bone remodeling, we developed a mouse model of HCS by introducing a pathogenic mutation (6272delT) into the murine Notch2 gene. By mCT and undecalcified histology, we observed generalized osteopenia in two independent mouse lines derived by injection of different targeted embryonic stem (ES) cell clones, yet acro-osteolysis did not occur until the age of 52 weeks. Cellular and dynamic histomorphometry revealed a high bone turnover situation in Notch2 þ/HCS mice, since osteoblast and osteoclast indices were significantly increased compared with wild-type littermates. Whereas ex vivo cultures failed to uncover cell-autonomous gain-of-functions within the osteoclast or osteoblast lineage, an unbiased RNA sequencing approach identified Tnfsf11 and Il6 as Notch-signaling target genes in bone marrow cells cultured under osteogenic conditions. Because we further observed that the high-turnover pathology of Notch2 þ/HCS mice was fully normalized by alendronate treatment, our results demonstrate that mutational activation of Notch2 does not directly control osteoblast activity but favors a pro-osteoclastic gene expression pattern, which in turn triggers high bone turnover.

Notch in skeletal physiology and disease

“…The mechanism involves a direct effect of Notch2 on osteoclast precursors as well as the induction of RANKL by cells of the osteoblast lineage. In an alternate murine model of HCS, an increased expression of interleukin 6 was reported in bone marrow cells, and this cytokine could contribute to the enhanced bone resorption observed [40]. Notch2 tm1.1Ecan mutant mice are sensitized to the development of osteoarthritis, a mechanism that may also involve an enhanced expression of interleukin 6 by the chondrocyte of HCS mutant mice [41].…”

NOTCH2 Hajdu-Cheney Mutations Escape SCFFBW7-Dependent Proteolysis to Promote Osteoporosis

“…In recent mouse studies, LyzM-Cre mediated myeloid-specific Notch1 −/− Notch2 −/− Notch3 −/− triple knockout facilitates osteoblast formation and concomitantly impairs osteoclastogenesis (Bai et al, 2008). Other recent studies demonstrated that the Notch2 HCS mutant mice faithfully recapitulate the features of HCS (Canalis et al, 2016; Vollersen et al, 2017). Additionally, a recent case study reported that the administration of denosumab, a RANKL antagonist, increases bone mineral density in a patient with HCS (Adami et al, 2016).…”

“…HCS is an ideal model to study a physiological function of NOTCH2 as C-terminal truncation mutations in HCS are shown to confer gain-of-function properties to Notch2 in mice (Canalis et al, 2017; Canalis et al, 2016; Vollersen et al, 2017). Moreover, most identified HCS NOTCH2 mutations cluster between the second nuclear localization signal (NLS) and the PEST domain, resulting in C-terminal premature truncation of NOTCH2 that lacks the PEST sequence (Figure 3A).…”

“…Recent studies indicate that NOTCH2-dependent osteoclastic bone resorption results in HCS development (Canalis et al, 2017; Canalis et al, 2016; Isidor et al, 2011; Simpson et al, 2011; Vollersen et al, 2017). Notably, the administration of anti-NOTCH2 NRR antibody in the HCS mouse reverses the osteopenia phenotype (Canalis et al, 2017).…”

Induction of the Hajdu-Cheney Syndrome Mutation in CD19 B Cells in Mice Alters B-Cell Allocation but Not Skeletal Homeostasis

“…Our previous work demonstrated that Notch2 DPEST/DPEST osteoblasts express increased levels of Tnfsf11 mRNA, suggesting that osteoblast-derived RANKL is responsible for the enhanced bone resorption in Notch2 tm1.1ECan mutant mice. 35,52,57 Similarly, a subject with HCS and severe osteoporosis was reported to present with elevated levels of RANKL in the serum. 18 Notch2 tm1.1ECan and control sex-matched littermates were subjected to splenectomies or sham interventions at 1 month of age and sacrificed 1 month later for analysis.…”