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. 2009 Dec 22;107(2):906–911. doi: 10.1073/pnas.0913415107

Fig. 4.

Fig. 4.

(A) PLC-γ1 is part of the protein complex that associates with TonEBP/OREBP at its DNA binding sites. EMSA binding reactions were performed with nuclear extracts of HEK293 cells exposed to 500 mosmol/kg medium (NaCl added) for 2 h. The ORE probe (bp −1,238 to −1,104 of the human AR gene) contains three OREs. Additions to reactions include antibodies that recognize PLC-γ1 and TonEBP/OREBP or 10-fold excess of nonbiotinylated probe (representative of three independent experiments). (B and C) PLC-γ1 contributes to high NaCl–dependent TonEBP/OREBP transactivating activity. (B) PLC-γ1 Null and Plus cells were cotransfected with pFR-Luc and TonEBP/OREBP C-terminal GAL4dbd-548-1531. After 24 h, osmolality either was kept at 300 mosmol/kg or was increased to 500 mosmol/kg for 16 h by adding NaCl. (C) HEK293 cells were cotransfected with pFR-Luc and TonEBP/OREBP C-terminal GAL4dbd-548-1531 and with either wild-type PLC-γ1 (WT), PLC-γ1 in which the SH3 domain was deleted (ΔSH3), or with PLC-γ1 in which the SH3 and both SH2 domains were deleted (ΔSH3/SH2-N&C). After 24 h, osmolality either was kept at 300 mosmol/kg or was increased to 500 mosmol/kg for 16 h by adding NaCl. Mean ± SEM. *P ≤ 0.05; n = 3.