ROLE OF AHR/ARNT SIGNALING ON MATRIX REMODELING IN SKIN

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DESCRIPTION (Adapted from the Candidate's Abstract) TCDD exposure leads to severe skin lesions known as chloracne. These lesions are characterized by hyperproliferation, hyperdifferentiation, and abnormal migration of keratinocytes in the stratified squamous epithelia. It is unclear how TCDD exposure leads to the skin pathology. Most of the studies on the molecular mechanisms of TCDD function have focused on the ligand-activated transcription factor AhR/Arnt. Binding of TCDD to AhR initiates heterodimerization with Arnt, and ultimately alters transcription through binding of this complex to specific sites (XRES) in the 5' regions of AhR responsive genes. TCDD target genes identified include the Phase I and Phase II detoxifying enzymes, as well as cytokines (TGF-B, IL-1B), transcription factors (Fos, Jun), and an extracellular matrix remodeling inhibitory proteins (PAI-2). Metabolism of the extracellular matrix is required in situations that require tissue remodeling or cell migration, such as wound healing. The majority of the matrix degradation is accomplished through the activity of members of the matrix metalloproteinase (MMP) family. A transcription factor database search identified two potential XRE sites in the 5' region of MMP-1, and preliminary results demonstrate that MMP-1 MRNA levels increase upon TCDD treatment of keratinocytes. This data suggests a link between TCDD/AhR/Arnt and matrix remodeling. The candidate is currently a post-doctoral fellow in the laboratory of Dr. B. L. Allen-Hoffmann at the University of Wisconsin- Madison, where her project is to generate a tissue specific knockout mouse of AhR in stratified squamous epithelia to study the effects of AhR loss on skin structure and differentiation. The goal of this proposal is to study the skin pathology resulting from TCDD exposure in intact tissue, and determine the role of the AhR/Arnt pathway on MMPs and their inhibitors, the TIMPS, into the affected skin. Therefore the specific aims of this proposal are: (1) create a mouse model, using the Cre/loxP recombination system of bacteriophage P1, to conditionally knock out the Arnt gene in the replicating keratinocytes of murine skin. Use this mouse model, along with the AhR tissue specific knockout, to study the effect of AhR and Arnt loss on MMP/TIMP expression and TCDD skin pathologies. (2) Use keratinocytes in an organotypic cell culture system to study the effect of TCDD on tissue architecture and gene expression of MMPs/TIMPS.