Heterozygous inactivation of Gnas in adipose-derived mesenchymal progenitor cells enhances osteoblast differentiation and promotes heterotopic ossification

摘要

Human genetic disorders sharing the common feature of subcutaneous heterotopic ossification (HO) are caused by heterozygous inactivating mutations in GNAS, a gene encoding multiple transcripts including two stimulatory G-proteins, the α-subunit of the stimulatory G-protein (Gsα) of adenylyl cyclase and the ‘extra-long” form of Gsα, XLαs. In one such disorder, progressive osseous heteroplasia (POH), bone formation initiates within subcutaneous fat before progressing to deeper tissues, suggesting that osteogenesis may involve abnormal differentiation of mesenchymal precursors that are present in adipose tissues. We determined by immunohistochemical analysis that GNAS protein expression is limited to Gsα in bone-lining cells and to Gsα and XLαs in osteocytes. By contrast, the GNAS proteins Gsα, XLαs, and NESP55 are detected in adipocytes and in adipose stroma. Although Gnas transcripts, as assessed by qRT-PCR, show no significant changes upon osteoblast differentiation of bone-derived precursor cells, the abundance of these transcripts is enhanced by osteoblast differentiation of adipose-derived mesenchymal progenitors. Using a mouse knockout model, we determined that heterozygous inactivation of Gnas (by disruption of the Gsα-specific exon 1) abrogates upregulation of multiple Gnas transcripts that normally occurs with osteoblast differentiation in wild-type adipose stromal cells. These transcriptional changes in Gnas^+/− mice are accompanied by accelerated osteoblast differentiation of adipose stromal cells in vitro. In vivo, altered osteoblast differentiation in Gnas^+/− mice manifests as subcutaneous HO by an intramembranous process. Taken together, these data suggest that Gnas is a key regulator of fate decisions in adipose-derived mesenchymal progenitor cells, specifically those that are involved in bone formation.