Background/Purpose: The enzyme tartrate-resistant acid phosphatase (TRAP) is highly expressed in osteoclasts. One of the main substrates for TRAP in bone matrix is osteopontin (OPN).  Biallelic mutations in the gene, ACP5, that encodes TRAP, result in an immuno-osseous disease called spondyloenchondrodysplasia (SPENCD).  In addition to bone and neurological abnormalities, SPENCD patients also develop autoimmune disorders such as systemic lupus erythematosus (SLE). Of note, SPENCD patients demonstrate evidence of increased interferon-alpha (IFN-α) production in their blood. Since very little is known about the function of TRAP in immune cells, the objectives of our study were to determine whether OPN is a substrate for TRAP and to define the consequences of TRAP deficiency in immune cells.  

Methods: Co-localization of TRAP and OPN was determined by confocal microscopy and also by immunoprecipitation and western blot analysis (IP-western).  TRAP overexpression or knockdown was performed by transfection with cDNA or shRNA, respectively.  Expression of IFN-α and IFN signature genes (ISGs) were determined by enzyme-linked immunosorbent assay (ELISA) and quantitative PCR (qPCR).  Phosphatase activity was quantified by Biomol Green fluorimetry.

Results: We observed that TRAP co-localized with OPN in early endosomes and the Golgi in both primary macrophages as well as in plasmacytoid dendritic cells (pDC).  Co-localization was confirmed biochemically: following co-transfection of cDNAs encoding TRAP and OPN in 293 cells, we reciprocally co-immunoprecipitated TRAP and OPN as determined by western blots. Also, in macrophages, anti-TRAP antibody immunoprecipitated both TRAP and OPN, indicating that they interacted with each other in primary non-transfected cells.  To confirm that OPN was indeed a substrate for TRAP, we observed that recombinant human TRAP dephosphorylated OPN by the release of free phosphate in an in vitroassay.  To relate the functional significance of TRAP deficiency to IFN-α production, we knocked down the expression of TRAP in pDC.  We observed that TRAP specific shRNA, but not scrambled shRNA, increased the expression of IFN-α and IFN signature genes (ISGs). 

Conclusion: Taken together, these findings indicate that TRAP and OPN co-localize and that OPN is a substrate for TRAP in immune cells.  Significantly, TRAP deficiency in pDC leads to increased IFN-α production providing an explanation for why TRAP mutations lead to a lupus-like disease in SPENCD patients.

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/tartrate-resistant-acid-phosphatase-deficiency-in-the-predisposition-to-systemic-lupus-erythematosus/