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Mechanical stimulation of polycystin-1 induces human osteoblastic gene expression via potentiation of the calcineurin/NFAT signaling axis

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Abstract

Mechanical forces trigger biological responses in bone cells that ultimately control osteoblastogenesis and bone program. Although several mechanosensors have been postulated, the precise mechanotransduction pathway remains obscure as the initial mechanosensing event has not yet been identified. Studies in kidney cells have shown that polycystin-1 (PC1), via its extracellular N-terminal part, may function as an “antenna-like” protein providing a linkage between environmental cues and their conversion into biochemical responses that regulate various cellular processes via the calcineurin/NFAT pathway. Here we explored the involvement of PC1 in mechanical load (stretching)-induced signaling cascades that control osteoblastogenesis/bone formation. FACS and TransAM Transcription Factor ELISA analyses employing extracts from primary human osteoblast-like, PC1 expressing cells subjected to mechanical stretching (0–6 h) revealed that unphosphorylated/DNA-binding competent NFATc1 increased at 0.5–1 h and returned to normal at 6 h. In accordance with the activation mechanism of NFATc1, stretching of cultured cells pre-treated with cyclosporin A (CsA, a specific inhibitor of the calcineurin/NFAT pathway) abrogated the observed decrease in the abundance of the cytoplasmic pNFATc1 (phosphorylated/inactive) species. Furthermore, stretching of osteoblastic cells pre-treated with an antibody against the mechanosensing N-terminal part of PC1 also abrogated the observed decrease in the cytoplasmic levels of the inactive pNFATc1 species. Importantly, under similar conditions (pre-incubation of stretched cells with the inhibitory anti-PC1 antibody), the expression of the key osteoblastic, NFATc1-target gene runx2 decreased compared to untreated cells. Therefore, PC1 acts as chief mechanosensing molecule that modulates osteoblastic gene transcription and hence bone-cell differentiation through the calcineurin/NFAT signaling cascade.

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Abbreviations

ADPKD:

Autosomal dominant polycystic kidney disease

CsA:

Cyclosporin A

FACS:

Fluorescent-activated cell sorting

hPDL cells:

Human periodontal ligament cells

NFAT:

Nuclear factor of activated T cells

NFATc1:

Nuclear factor of activated T cells, cytoplasmic 1

PC1:

Polycystin-1

PC2:

Polycystin-2

Pkd ½ :

Polycystic kidney disease 1/2 (polycystin 1/2) gene

pNFAT:

Phosphorylated nuclear factor of activated T cells

Runx2:

Runt-related transcription factor 2

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Acknowledgments

We are grateful to Dr. G.G. Germino for the anti-CT antibody and the stably transfected MDCK-PKD1 cell line. We thank Dr. Oxana Ibraghimov-Beskrovnaya for the inhibitory anti-Ig-PKD1 antibody.

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Authors and Affiliations

  1. Cellular and Molecular Biomechanics Unit, Department of Biological Chemistry, University of Athens Medical School, 75, M. Asias Street, 11527, Athens, Greece

    Georgia Dalagiorgou, Christina Piperi, Christos Adamopoulos, Efthimia K. Basdra & Athanasios G. Papavassiliou

  2. Molecular Virology Laboratory, Hellenic Pasteur Institute, 11527, Athens, Greece

    Urania Georgopoulou

Authors
  1. Georgia Dalagiorgou
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  2. Christina Piperi
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  3. Urania Georgopoulou
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  4. Christos Adamopoulos
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  5. Efthimia K. Basdra
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  6. Athanasios G. Papavassiliou
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Corresponding author

Correspondence to Efthimia K. Basdra.

Additional information

G. Dalagiorgou and C. Piperi contributed equally to this study.

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Dalagiorgou, G., Piperi, C., Georgopoulou, U. et al. Mechanical stimulation of polycystin-1 induces human osteoblastic gene expression via potentiation of the calcineurin/NFAT signaling axis. Cell. Mol. Life Sci. 70, 167–180 (2013). https://doi.org/10.1007/s00018-012-1164-5

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  • DOI: https://doi.org/10.1007/s00018-012-1164-5

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