Abstract
Ca(2+) signaling is essential for bone homeostasis and skeletal development. Here, we show that the transient receptor potential canonical 1 (TRPC1) channel and the inhibitor of MyoD family, I-mfa, function antagonistically in the regulation of osteoclastogenesis. I-mfa null mice have an osteopenic phenotype characterized by increased osteoclast numbers and surface, which are normalized in mice lacking both Trpc1 and I-mfa. In vitro differentiation of pre-osteoclasts derived from I-mfa-deficient mice leads to an increased number of mature osteoclasts and higher bone resorption per osteoclast. These parameters return to normal levels in osteoclasts derived from double mutant mice. Consistently, whole cell currents activated in response to the depletion of intracellular Ca(2+) stores are larger in pre-osteoclasts derived from I-mfa knock-out mice compared with currents in wild type mice and normalized in cells derived from double mutant mice, suggesting a cell-autonomous effect of I-mfa on TRPC1 in these cells. A new splice variant of TRPC1 (TRPC1ε) was identified in early pre-osteoclasts. Heterologous expression of TRPC1ε in HEK293 cells revealed that it is unique among all known TRPC1 isoforms in its ability to amplify the activity of the Ca(2+) release-activated Ca(2+) (CRAC) channel, mediating store-operated currents. TRPC1ε physically interacts with Orai1, the pore-forming subunit of the CRAC channel, and I-mfa is recruited to the TRPC1ε-Orai1 complex through TRPC1ε suppressing CRAC channel activity. We propose that the positive and negative modulation of the CRAC channel by TRPC1ε and I-mfa, respectively, fine-tunes the dynamic range of the CRAC channel regulating osteoclastogenesis.
Highlights
Whole cell currents activated in response to the depletion of intracellular Ca2؉ stores are larger in pre-osteoclasts derived from I-mfa knock-out mice compared with currents in wild type mice and normalized in cells derived from double mutant mice, suggesting a cell-autonomous effect of I-mfa on transient receptor potential canonical 1 (TRPC1) in these cells
The higher osteoclast numbers and larger erosion surface per bone surface in I-mfa null mice were normalized in the double knock-out mice (Fig. 1, A–D), suggesting that the increased osteoclastogenesis seen in I-mfa mutant mice was related to increased activity of TRPC1
Our study provides several lines of evidence supporting the hypothesis that Trpc1 and I-mfa genetically and functionally interact to regulate osteoclastogenesis through store-operated Ca2ϩ entry channels
Summary
Mature osteoclasts are derived from hematopoietic stem cells through a series of events initiated by the formation of myeloid precursors in response to macrophage-colony stimulating factor (M-CSF)5 [1] These precursors differentiate into multinucleated osteoclasts in a multistep process dependent on M-CSF and receptor activator of nuclear factor-B ligand (RANKL) [2]. Both of these factors act through Ca2ϩ signaling to induce downstream regulators of osteoclastogenesis such as nuclear factor of activated T cells c1 (NFATc1), NF-B, c-fos, -catenin, and others [3, 4].
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