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. 2014 Oct 7;9(10):e108942. doi: 10.1371/journal.pone.0108942

Figure 3. Dependence of EMT reprogramming on recruitment of PI3K/p85 but not Grb2.

Figure 3

(A) Immunoprecipitation (IP) and immunoblot (IB) of NKG2D and/or DAP10 (N87Q* or M88Q*) in transduced MCF-10AT cells (top two panels). Bottom three panels show primary detection of DAP10 and the mutually exclusive association of its N87Q* and M88Q* variants with p85 and Grb2, respectively. (B) Signaling functionality of the DAP10 N87Q* and M88Q* variants. Note that ERK is downstream of PI3K. EGF was added for control activation, DMSO for solvent control. (C, D) Reversal of EMT protein marker and transcription marker profiles by expression of DAP10 (M88Q*) with impaired binding of p85, shown by immunoblot and/or RT-PCR. (E) DAP10 variant M88Q* but not N87Q* reduces migratory and invasive activities of Dox-induced MCF-10AT cells expressing the respective NKG2D–DAP10 mutant complexes. Bars represent mean cell numbers (+/− SD) derived from three independent in vitro migration and invasion experiments with each four microscopic field counts. Asterisks denote p<0.005.