Core binding factor beta (Cbfβ), the partner protein of Runx family transcription factors, enhances Runx function by increasing the binding of Runx to DNA. Null mutations of Cbfb result in embryonic death, which can be rescued by restoring fetal hematopoiesis but only until birth, where bone formation is still nearly absent. Here, we address a direct role of Cbfβ in skeletal homeostasis by generating osteoblast‐specific Cbfβ‐deficient mice (Cbfb^Δob/Δob) from Cbfb‐floxed mice crossed with mice expressing Cre from the Col1a1 promoter. Cbfb^Δob/Δob mice showed normal growth and development but exhibited reduced bone mass, particularly of cortical bone. The reduction of bone mass in Cbfb^Δob/Δob mice is similar to the phenotype of mice with haploinsufficiency of Runx2. Although the number of osteoblasts remained unchanged, the number of active osteoblasts decreased in Cbfb^Δob/Δob mice and resulted in lower mineral apposition rate. Immunohistochemical and quantitative real‐time PCR analyses showed that the expression of osteogenic markers, including Runx2, osterix, osteocalcin, and osteopontin, was significantly repressed in Cbfb^Δob/Δob mice compared with wild‐type mice. Cbfβ deficiency also reduced Runx2 protein levels in osteoblasts. The mechanism was revealed by forced expression of Cbfβ, which increased Runx2 protein levels in vitro by inhibiting polyubiquitination‐mediated proteosomal degradation. Collectively, these findings indicate that Cbfβ stabilizes Runx2 in osteoblasts by forming a complex and thus facilitates the proper maintenance of bone mass, particularly cortical bone. © 2015 American Society for Bone and Mineral Research.

The transdifferentiation of vascular smooth muscle cells (VSMCs) into osteoblast-like cells has been implicated in the context of vascular calcification. We investigated the roles of vitamin D receptor (Vdr) and runt-related transcription factor 2 (Runx2) in the osteoblastic differentiation of VSMCs in response to vitamin D₃ using in vitro VSMCs cultures and in vivo in Vdr knockout (Vdr^-/-) and Runx2 carboxy-terminus truncated heterozygous (Runx2^+/ΔC) mice. Treatment of VSMCs with active vitamin D₃ promoted matrix mineral deposition, and increased the expressions of Vdr, Runx2, and of osteoblastic genes but decreased the expression of smooth muscle myosin heavy chain in primary VSMCs cultures. Immunoprecipitation experiments suggested an interaction between Vdr and Runx2. Furthermore, silencing Vdr or Runx2 attenuated the procalcific effects of vitamin D₃. Functional cooperation between Vdr and Runx2 in vascular calcification was also confirmed in in vivo mouse models. Vascular calcification induced by high-dose vitamin D₃ was completely inhibited in Vdr^-/- or Runx2^+/ΔC mice, despite elevated levels of serum calcium or alkaline phosphatase. Collectively, these findings suggest that functional cooperation between Vdr and Runx2 is necessary for vascular calcification in response to vitamin D₃.

Tendon plays a critical role in the joint movement by transmitting force from muscle to bone. This transmission of force is facilitated by its specialized structure, which consists of highly aligned extracellular matrix consisting predominantly of type I collagen. Tenocytes, fibroblast-like tendon cells residing between the parallel collagen fibers, regulate this specialized tendon matrix. Despite the importance of collagen structure and tenocyte function, the biological mechanisms regulating fibrillogenesis and tenocyte maturation are not well understood. Here we examine the function of Reticulocalbin 3 (Rcn3) in collagen fibrillogenesis and tenocyte maturation during postnatal tendon development using a genetic mouse model. Loss of Rcn3 in tendon caused decreased tendon thickness, abnormal tendon cell maturation, and decreased mechanical properties. Interestingly, Rcn3 deficient mice exhibited a smaller collagen fibril distribution and over-hydroxylation in C-telopeptide cross-linking lysine from α1(1) chain. Additionally, the proline 3-hydroxylation sites in type I collagen were also over-hydroxylated in Rcn3 deficient mice. Our data collectively suggest that Rcn3 is a pivotal regulator of collagen fibrillogenesis and tenocyte maturation during postnatal tendon development.