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. Author manuscript; available in PMC: 2017 Nov 15.
Published in final edited form as: J Bone Miner Res. 2017 Aug 2;32(11):2207–2218. doi: 10.1002/jbmr.3205

Figure 5. DAP12 is required for M-CSF or RANKL-induced DOK3 phosphorylation.

Figure 5

(A) RAW264.7 cells were crosslinked with anti-TREM2 antibody for 2, 5, and 20 minutes. Cells were subjected to immunoprecipitation with antibodies against phosphorylated-DAP12 (pDAP12), DOK3, or phosphorylated-tyrosine (pTyr). Protein complexes were analyzed by Western blotting with antibodies specific for SHIP1, pDAP12, and DOK3. Whole cell lysates (WCL) were immunoblotted for phosphorylated-SHIP1 (pSHIP1), total SHIP1, and DOK3. (B) BMMs were stimulated with 100 ng/ml MCSF and lysates immunoprecipitated with anti-DOK3 and anti-pTyr antibodies. WCL or co-immunoprecipitated complexes were immunoblotted for SHIP1 and DOK3. (C and D) WT and DAP12-/- BMMs were stimulated with M-CSF (100 ng/ml). Cell lysates were immunoprecipitated with anti-DOK3 (C) and probed with pTyr antibody and DOK3 or anti-Grb2 (D) and immunoblotted for DOK3 and Grb2. Experiment was repeated three times.