ERK and p38 MAPK inhibition controls NF-E2 degradation and profibrotic signaling in renal proximal tubule cells

Aims: Transforming growth factor-beta (TGF-beta) mediates fibrotic manifestations of diabetic nephropathy. We demonstrated proteasomal degradation of anti-fibrotic protein, nuclear factor-erythroid derived 2 (NF-E2), in TGF-beta treated human renal proximal tubule (HK-11) cells and in diabetic mouse kidneys. The current study examined the role of mitogen-activated protein kinase (MAPK) pathways in mediating NF-E2 proteasomal degradation and stimulating profibrotic signaling in HK-11 cells. Main methods: HK-11 cells were pretreated with vehicle or appropriate proteasome and MAPK inhibitors, MG132 (0.5 mu M), SB203580 (1 mu M), PD98059 (25 mu M) and SP600125 (10 mu M), respectively, followed by treatment with/without TGF-beta (10 ng/ml, 24 h). Cell lysates and kidney homogenates from FVB and OVE26 mice treated with/without MG132 were immunoblotted with appropriate antibodies. pUse vector and pUse-NF-E2 cDNA were transfected in HK-11 cells and effects of TGF-beta on JNK MAPK phosphorylation (pJNK) was examined. Key findings: We demonstrated activation of p38, ERK, and JNK MAPK pathways in TGF-beta treated HK-11 cells. Dual p38 and ERK MAPK blockade prevented TGF-beta-induced pSer82Hsp27, fibronectin and connective tissue growth factor (CTGF) expression while preserving NF-E2 expression. Blockade of JNK MAPK inhibited TGF-beta-induced CTGF expression without preserving NF-E2 expression. MG132 treatment prevented TGF-beta-induced pJNK in HK-11 cells and in type 1 diabetic OVE26 mouse kidneys, demonstrating that TGF-beta-and diabetes-induced pJNK occurs downstream of proteasome activation. A direct role for NF-E2 in modulating pJNK acti-vation was demonstrated by NF-E2 over-expression. Significance: ERK and p38 MAPK promotes NF-E2 proteasomal degradation while proteasome activation pro-motes pJNK and profibrotic signaling in renal proximal tubule cells.