Smad6 and Smad7 are inhibitory Smads (I-Smads) with differential inhibitory effects on the regulation of the cellular signalings induced by TGF-beta superfamily. Here, we show that phorbol ester 12-O-tetradecanoylphorbol- 13 -acetate (TPA) down-regulates Smad6 mRNA expression and up-regulates Smad7 mRNA expression in IMR-90, a human lung fibroblast cell line. These regulations of I-Smads by TPA were suppressed by one PKC inhibitor (Go6983), but not by another (Go6976). TPA treatment had little effect on the phosphorylation of novel PKCs (PKCdelta and PKCepsilon), but specifically induced PKCmu phosphorylation, and this effect was inhibited by Go6983, but not by Go6976. Additionally, Go6983 but not Go6976 inhibited ERK- and JNK-phosphorylation as well as Smad7 promoter activity induced by TPA. NIEK inhibitor U0126 inhibited the down-regulation of Smad6 mRNA expression but not the up-regulation of Smad7 mRNA expression. In contrast, JNK inhibitor SP600125 had no such effects. Luciferase reporter analysis revealed that TPA did not induce NF-kappaB activation. In addition, TPA up-regulated Smad7 expression in the presence of NF-kappaB inhibitor TLCK. These findings indicate that TPA down-regulates Smad6 expression presumably via PKCmu-ERK-dependent pathway and up-regulates Smad7 expression via PKCmu-dependent mechanism(s) which need no MAPK and NF-kappaB activation. (C) 2004 Elsevier Inc. All rights reserved.