Amino acid transporters regulate bone formation

Bone development and homeostasis are governed by a number of developmental signals, transcription factors and cellular metabolism. This process is also dependent on the orchestration of multiple cell types including osteoblasts, chondrocytes, skeletal stem cells and osteoclasts. Osteoblasts are the principal bone forming cells responsible for producing and secreting the type I collagen rich extracellular bone matrix. Protein synthesis is an energetically and biosynthetically demanding process. This requires copious amounts of ATP and amino acids amongst other metabolites. However, the precise mechanisms and systems that osteoblasts utilize to meet these synthetic demands are poorly understood. Previous studies have shown amino acid consumption is increased in osteoblasts during differentiation. This process is regulated by transcription factors ATF4 and FOXO. Additionally, osteogenic signals like WNT and PTH can stimulate amino acid uptake. For example, WNT signaling can rapidly stimulate glutamine uptake and metabolism required for osteoblast differentiation. Unfortunately, transporters mediating glutamine uptake in osteoblasts are unknown. Moreover, the mechanism by which WNT stimulates increased glutamine consumption is also unknown. We identified two amino acid transporters, Slc7a7 and Slc1a5, as the primary glutamine transporters in response to WNT. Slc7a7 is responsible for the rapid WNT-induced glutamine uptake via the -catenin dependent pathway. Conversely, Slc1a5 sustains basal glutamine uptake, which is regulated by ATF4 downstream of the mTORC1 pathway. In summary, these data demonstrate the biphasic role of WNT signaling in regulating glutamine consumption, by two amino acid transporters Slc7a7 and Slc1a5, during osteoblast differentiation. While we have shown the importance of glutamine in bone cells, the role of other amino acids is not clear. Proline has long been considered as a critical amino acid due to its enrichment in collagens. Furthermore, PTH stimulates proline consumption in osteoblasts. The transport of proline is characterized by its dependency on sodium and sensitivity to MEAIB. However, the precise transport system responsible for proline import is not known. Here we identified the amino acid transporter Slc38a2, which encodes SNAT2, as the primary proline transporter in osteoblasts. Deletion of Slc38a2 results in defects in both intramembranous and endochondral ossifications. The phenotype is associated with defective osteoblast differentiation highlighted by reduction of proline enriched proteins (e.g. RUNX2, OSX and COL1A1). Slc38a2 provides proline to support osteoblast differentiation through two mechanisms. First, majority of proline is directly incorporated into proteins and does not contribute to amino acid biosynthesis. Second, proline oxidation regulates bioenergetics required for osteoblast differentiation. These findings highlight the multifaceted functions of proline, which is provided by Slc38a2, in osteoblast differentiation and bone formation. Collectively, my work demonstrates the critical role of amino acid transporters in osteoblast differentiation and provides novel insights in their potential applications in treatments of bone diseases like osteoporosis and bone fracture.