Molecular mechanisms of differential transcriptional activity of AP-1 factors c-Jun, Fra-1 and ATF-2, and their functional significance in malignant melanoma

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In recent decades, many altered pathways regulating the development and progression of melanoma and the high migratory and invasive potential of melanoma cells have been identified, but a detailed understanding of defined molecular mechanisms in disease development and progression is still incomplete. A crucial step in the development and progression of melanoma is the de-regulation of tumor-promoting transcription factors, for example of the Activating Protein-1 (AP-1). AP-1 proteins regulate the expression of specific target genes, which leads to the de-regulation of tumor-relevant signaling pathways. Thus, AP-1 transcription factors play an important role in various tumor types, including malignant melanoma. A key feature of AP-1 complexes in the cell is their heterogeneity with respect to their dimer composition. Various studies have shown that differences in AP-1 dimer compositions cause a change in the selection of specific binding sites. However, it remains unclear which direct target genes of AP-1 homodimers or heterodimers cause the functional effects that support melanomagenesis.The AP-1 family member c-Jun is a major regulator of melanoma progression and acts by regulating target genes that support tumor cell proliferation and migration, thereby promoting the development of a malignant phenotype. The de-regulation of c-Jun, Fra-1 and ATF-2 are major events in melanoma development, but the functional relevance of their interplay and molecular effects on target gene expression could not be shown in detail so far.In the proposed project, the relevance of interaction of selected AP-1 transcription factors (c-Jun, Fra-1, and ATF-2) for the development and progression of malignant melanoma will be investigated.C-Jun, ATF-2 and Fra-1 ChIP-Seq data analysis will be used to analyze the DNA-binding sites of the individual transcription factors and to identify potentially different DNA-binding modalities of the dimers, which will be validated in further experiments (e.g. Luciferase reporter gene assay, Electrophoretic Mobility Shift Assay). Moreover, motifs near the identified AP-1 binding motifs will be defined and the influence of the presence or absence of AP-1 co-factors on regulatory mechanisms will be determined. In expression analyses (qRT-PCR, Western Blots) of the target genes confirmed by ChIP- and RNA-Seq in different melanoma cell lines versus melanocytes, possible de-regulations will be revealed. Target genes, which are directly regulated by c-Jun, Fra-1, and/or ATF-2 and show a de-regulation in melanoma cells compared to normal melanocytes, will be investigated using functional experiments (proliferation, migration, invasion assays, etc.) for their relevance and function in melanoma.The results of this project will make a significant contribution to elucidate of the role of AP-1 transcription factors in the development of melanoma and would also lead to the identification and development of new therapeutic targets and options.