Figure 2. LZK signals through endogenous MKK4-JNKs and JNK inhibition reduces LZK protein levels in N2a cells.

(A) N2a and HeLa cells were transfected with empty vector (EV), FLAG-LZK, or FLAG-LZK-K195A. HeLa cells collected 45 min after UV irradiation at 45 J/m² served as positive control for activation of endogenous JNK1/2, p38, and MKK4. Total cell lysates were immunoblotted for the indicated proteins. p-JNK indicating phospho-(Thr183/Tyr185)-JNK1/2; p-MKK4 indicating phospho-(Ser257)-MKK4. (B) Total lysates from N2a cells transfected with empty vector (EV), FLAG-LZK, or FLAG-LZK-K195A were immunoblotted for the indicated proteins. p-P38 indicates phospho-(Thr180/Tyr182)-p38; p-ERK1/2 indicates phospho-(Thr202/Tyr204)-ERK1/2. (C) N2a cells were transfected with empty vector (EV) or FLAG-LZK and treated with SP600125 at the indicated doses at the time of transfection. Total cell lysates were immunoblotted for the indicated proteins. p-cJun indicates phospho-(Ser63)-c-Jun. (D) Total lysates of N2a cells overexpressing empty vector (EV), DLK, LZK, or LZK-K195A were immunoblotted for the indicated proteins. (E) Total lysates of N2a cells overexpressing empty vector (EV), DLK, or LZK were immunoblotted for the indicated proteins.