Thursday, April 3, 2025
4/3/2025
Effects of TP63 mutations on keratinocyte adhesion and differentiation - PROJECT SUMMARY Ankyloblepharon ectodermal dysplasia and cleft lip/palate (AEC) and ectrodactyly ectodermal dysplasia and cleft lip/palate (EEC) are two ectodermal dysplasias. AEC and EEC are caused by mutations in the transcription factor TP63. TP63 is known to be essential for the development and maintenance of the epidermis and epidermal appendages. TP63 mutations in AEC and EEC (TP63-AEC and TP63-EEC) lead to epidermal phenotypes, including skin fragility. To develop therapeutic options, it is essential to understand the pathological mechanisms leading to these devastating manifestations. My preliminary data suggest that different mechanisms contribute to TP63-AEC and TP63-EEC skin pathology. TP63-AEC keratinocytes downregulate integrins required for extracellular matrix (ECM) adhesion while TP63-EEC keratinocytes do not. Further, TP63-AEC keratinocytes also show reduced ECM adhesion and migration, suggesting a possible link between integrin defects and abnormal keratinocyte behavior. TP63-AEC keratinocytes also demonstrated reduced epidermal differentiation while TP63-EEC keratinocytes displayed accelerated differentiation. Based on these observations, I hypothesize that adhesion and differentiation defects contribute to the epidermal phenotype observed in AEC and EEC patients. In order to study the molecular pathology underlying AEC and EEC, I will generate human NTERT keratinocyte lines expressing TP63-AEC and TP63-EEC mutant proteins. NTERT keratinocytes display normal keratinocyte behavior and can differentiate in 2D and 3D systems. Aim 1 will establish a mechanistic link between specific integrins and their contribution to TP63-AEC keratinocyte adhesion and migration defects. Aim 2 will address the apparent different mechanisms between TP63-AEC and TP63-EEC. I will generate epidermal equivalents expressing TP63-AEC or TP63-EEC proteins and use immunofluorescent staining and single-cell RNA sequencing (scRNAseq) to identify differences in various subpopulations of differentiating keratinocytes. The current proposal aims to determine the mechanistic consequences of TP63- AEC and TP63-EEC mutations on keratinocyte ECM adhesion and epidermal differentiation. This work has the potential to lead to the identification of novel disease pathways, and might lead to the discovery of novel therapeutic targets.
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