Ricerc@Sapienza

FGFR2b is the FGFR2 epithelial splicing variant whose signaling triggers different outcomes in epithelial cells, including differentiation. Starting from the evidence that several PKC isoforms participate in the regulation of cell differentiation, in this project we propose to assess if the well-known role of FGFR2b in early steps of keratinocyte differentiation could involve PKCdelta pathway. At the same time we also propose to clarify if this receptor can also control intermediate differentiation always via PKCdelta and/or late differentiation through PKCalpha. Using 2D cultures or 3D skin equivalents prepared with keratinocytes stably overexpressing or not FGFR2b, we propose to analyze the expression and the distribution throughout the epidermal layers of early, intermediate and late differentiation markers by molecular, immunofluorescence and biochemical approaches, while the ultrastructural aspects of skin equivalent differentiation will be checked in detail by transmission electron microscopy. Then, the possible differential contribution of PKCdelta and PKCalpha in FGFR2b-induced differentiation will be investigated by the use of isoform specific inhibitors of PKC activity and definitely confirmed using silencing approaches. Because it is well known that, in keratinocytes, PKCs isoforms control early/intermediate and late differentiation through a functional crosstalk with KLF4 and DLX3 respectively, the possible ability of FGFR2b to modulate these two transcription factors in both 2D cultures and 3D rafts of the different clones will be also investigated by molecular approaches.
The expected results will make it possible to advance in the knowledge of the complex signaling network through which FGFRs might control keratinocyte differentiation. These networks involving PKCs/transcription factors axis underlies the physiological progression along the differentiation program, and could be unbalanced in several differentiation disorders, including SCCs.