The TGF?1 pathway is required for NF?B dependent gene expression in mouse keratinocytes.

ABSTRACT: The transforming growth factor-beta 1 (TGF?1) and NF?B pathways are important regulators of epidermal homeostasis, inflammatory responses and carcinogenesis. Previous studies have shown extensive crosstalk between these pathways that is cell type and context dependent, but this has not been well-characterized in epidermal keratinocytes. Here we show that in primary mouse keratinocytes, TGF?1 induces NF?B-luciferase reporter activity that is dependent on both NF?B and Smad3. TGF?1-induced NF?B-luciferase activity was blocked by the I?B inhibitor parthenolide, the I?B super-repressor, a dominant negative TGF?1-activated kinase 1 (TAK1) and genetic deletion of NF?B1. Coexpression of NF?B p50 or p65 subunits enhanced NF?B-luciferase activity. Similarly, inhibition of the TGF?1 type I receptor with SB431542 or genetic deletion of Smad3 blocked TGF?1 induction of NF?B-luciferase. TGF?1 rapidly induced IKK phosphorylation but did not cause a detectable decrease in cytoplasmic I?B levels or nuclear translocation of NF?B subunits, although EMSA showed rapid NF?B nuclear binding activity that could be blocked by SB431542 treatment. TNF?, a well characterized NF?B target gene was also induced by TGF?1 and this was blocked in NF?B+/- and -/- keratinocytes and by the I?B super-repressor. To test the effects of the TGF?1 pathway on a biologically relevant activator of NF?B, we exposed mice and primary keratinocytes in culture to UVB irradiation. In primary keratinocytes UVB caused a detectable increase in levels of Smad2 phosphorylation that was dependent on ALK5, but no significant increase in SBE-dependent gene expression. Inhibition of TGF?1 signaling in primary keratinocytes with SB431542 or genetic deletion of Tgfb1 or Smad3 suppressed UVB induction of TNF? message. Similarly, UVB induction of TNF? mRNA was blocked in skin of Tgfb1+/- mice. These studies demonstrate that intact TGF?1 signaling is required for NF?B-dependent gene expression in mouse keratinocytes and skin and suggest that a convergence of these pathways in the nucleus rather than the cytoplasm may be critical for regulation of inflammatory pathways in skin by TGF?1.

SUBMITTER: Hogan KA 

PROVIDER: S-EPMC3942663 | biostudies-literature | 2013 Dec

REPOSITORIES: biostudies-literature