MODULATION OF NUCLEAR FACTOR KAPPA B (NFκB) TRANSACTIVATION BY TRANSFORMING GROWTH FACTOR β-1 (TGFβ-1) IN KERATINOCYTES: IMPLICATIONS FOR RESPONSIVENESS TO ULTRAVIOLET RADIATION (UVB)
Open Access
- Author:
- Hogan, Kelly A
- Graduate Program:
- Integrative Biosciences
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- October 04, 2011
- Committee Members:
- Adam Glick Ph D, Dissertation Advisor/Co-Advisor
Adam Bleier Glick, Committee Chair/Co-Chair
John Patrick Vanden Heuvel, Committee Member
Kumble Sandeep Prabhu, Committee Member
Andrea Marie Mastro, Committee Member
Avery August, Committee Member
Peter John Hudson, Committee Member - Keywords:
- TGF beta-1
ultraviolet radiation
NF kappa B
keratinocytes
TNF alpha
skin - Abstract:
- Molecular crosstalk leading to the integration of signal transduction pathways—and the formation of a signaling network—is particularly important for maintaining cellular homeostasis. Stimuli received at the cell surface and transduced to the nucleus can expect to be modified by any number of inputs in a highly context-dependent and cell-specific manner. Nuclear factor kappa B (NFκB) and transforming growth factor β-1 (TGFβ-1) are not only critical factors mediating inflammation, but they also play a substantial role in cancer progression. Therefore, understanding the intersection of these factors may shed light on inflammatory diseases and progression of cancer in skin. However, little is known about how TGFβ-1 and NFκB interact in keratinocytes, which rely heavily on both factors to maintain homeostasis. These studies provide data in keratinocytes that suggest TGFβ-1 modulated NFκB-dependent expression of proinflammatory cytokines, namely TNFα. Although results in these studies fail to show TGFβ-1-mediated activation of upstream molecules of the canonical NFκB pathway or translocation of NFκB, preliminary evidence reveals that TGFβ-1 activating kinase (TAK-1) may provide a molecular link between TGFβ-1 receptor activation and NFκB transactivation. In spite of the fact that upstream signaling events are only speculative and part of ongoing inquiry, results presented in this chapter support the hypothesis that TGFβ-1-mediated NFκB transactivation of gene expression is Smad3-dependent. Furthermore, TGFβ-1 potentiates NFκB binding to consensus DNA sites, which putatively involves both the p50 and p65 subunit. The biological relevance of TGFβ-1 and NFκB crosstalk leading to expression of proinflammatory cytokines is also explored. Preliminary evidence suggests that this pathway may have a role in TGFβ-1-mediated apoptosis, differentiation, and ras-mediated induction of NFκB-dependent genes in keratinocytes. These studies are the first to show an intersection between TGFβ-1 and NFκB pathways, which may represent a mechanism by which TGFβ-1 ‘tunes’ or modulates NFκB-dependent gene expression. The biological relevance of TGFβ-1-mediated TNFα was then explored in the context of ultraviolet radiation responsiveness, which elicits an inflammatory response involving the proinflammatory cytokine TNFα. Ultraviolet radiation, particularly the UVB wavelengths ranging from 280-320 nm, is a whole carcinogen capable of initiating and promoting squamous cell carcinoma (SCC), among other types of skin cancer, in both humans and laboratory rodents after repeated UVB exposure over time. Responsiveness to UVB, specifically, has not been particularly well-characterized in keratinocytes. Presently, the literature reflects more rigorous characterization of the UVC wavelengths in cell types that are typically not sun-exposed. Furthermore, published studies using the mouse as a model to inquire into TGFβ-1-mediated UVB responsiveness are non-existent. The present studies, performed in mouse and in primary keratinocytes isolated from mouse, demonstrate the intersection of TGFβ-1 and NFκB in the context of UVB responsiveness. Specifically, the hypothesis to be tested predicts that response to UVB will be partially dependent on TGFβ-1 signaling. UVB treated mice and keratinocytes in culture demonstrated Smad3- and NFκB-dependent expression of the proinflammatory cytokine TNFα between 2-6 h post treatment and required an intact TGFβ-1 signaling pathway. Furthermore, an acute decrement in Smad7 expression was observed initially, but restored to control levels by 6 h. Smad7 repression also appears to be partially Smad3 dependent. The results of these studies also demonstrated for the first time TGFβ-1-mediated NFκB binding of p50 and p65 subunits to DNA following UVB exposure. Although degradation of IκB or translocation of the p50 subunit was not observed, the data presented herein suggested a scenario whereby the NFκB-dependent proinflammatory cytokine was expressed in a p50- and Smad3-dependent manner. Taken together, it is likely that TGFβ-1 is among the pathways involving NFκB transactivation that modulate or tune response to UVB.