Hairy and Enhancer of Split-related with YRPW Motif (HEY)2 Regulates Bone Remodeling in Mice

Notch induces Hairy and Enhancer of Split-related with YRPW motif (Hey)1, Hey2, and HeyL expression in osteoblasts, but the contributions of these genes to the skeletal effects of Notch are not fully understood. HEY1 misexpression has limited skeletal impact, female HeyL null mice display increased bone mass, and Hey2 inactivation is developmentally lethal. To inactivate Hey2 in immature or mature osteoblasts, Hey2(loxP/loxP) mice were crossed with transgenics expressing CRE under the control of the osterix (Osx-Cre) or osteocalcin (Oc-Cre) promoters to generate Osx-Cre(+/-);Hey2(/) or Oc-Cre(+/-);Hey2(/) mice. Trabecular bone volume increased in 3-month-old Osx-Cre(+/-);Hey2(/) and Oc-Cre(+/-);Hey2(/) male mice and in 1-month-old Oc-Cre(+/-);Hey2(/) female mice, although 3-month-old Oc-Cre(+/-);Hey2(/) females developed osteopenia. Alkaline phosphatase liver/bone/kidney (ALPL) expression and activity were suppressed in osteoblasts from Oc-Cre(+/-);Hey2(/) mice of both sexes. To overexpress HEY2 in osteoblasts, transgenic mice where a 3.6-kb fragment of the rat collagen type-I 1 promoter directs HEY2 expression were created. Three-month-old Hey2 transgenic males exhibited decreased osteoblast activity and increased bone resorption and developed osteopenia at 6 months of age. Hey2 transgenic females exhibited reduced osteoblast number and function, but no changes in bone resorption. HEY2 overexpression in osteoblasts from mice of both sexes inhibited ALPL expression and activity and suppressed osteocalcin transcripts in cells from male mice only. HEY2 overexpression in osteoblasts from male mice enhanced bone resorption by co-cultured splenocytes and induced interleukin-6, a molecule that promotes osteoclastogenesis. In conclusion, HEY2 decreases skeletal mass and regulates bone remodeling in male mice.