Skip to main content
. 2018 Sep 14;293(44):17229–17239. doi: 10.1074/jbc.RA118.003926

Figure 4.

Figure 4.

JNK1 activation induces mSin1 expression and mTORC2 stabilization. A, nonfibrotic MCs (n = 6) were infected with a lentivirus containing an empty vector or a vector that expresses constitutively active JNK1 tagged with FLAG and then treated with the mTOR inhibitor AZD8055 (250 nm for 24 h). Cell lysates were immunoblotted against mSin1, collagen I, GAPDH (loading control), FLAG, and the phosphorylated and total forms of Jun (Ser-73), AKT (Ser-473), p70S6K1(Thr-389), 4E-BP1 (Thr-37/46), and mTOR (Ser-2481). B, nonfibrotic MCs (n = 6) were infected with a lentivirus containing an empty vector or vector that expresses constitutively active JNK1, followed by transfection with scrambled or mSIN1-specific siRNA. Cell lysates were immunoblotted with antibodies as described in A. C, nonfibrotic MCs (n = 8) were pretreated with 10 μm JNK inhibitor (SP600125) and then stimulated with LPA (10 μm for 6 h). Cell lysates were immunoblotted with antibodies against mSin1, collagen I, and phosphorylated and total forms of JNK1/2 (Thr-183/Tyr-185), AKT (Ser-473), and mTOR (Ser-2481) as well as rictor, raptor, and GAPDH. Cell lysates were immunoprecipitated using antibodies against mSin1, rictor, or mTOR. Each of the immunoprecipitates was analyzed for rictor, mSin1, phosphorylated mTOR (Ser-2481), and total mTOR. Representative blots are shown for each experiment.