Deciphering transcriptional regulation of NF-kB target genes using integrative omics approaches

Projektbeschreibung

Dem Mechanismus der transkriptionellen Aktivierung des Transkriptionsfaktors NF-kB auf der Spur

Transkriptionsfaktoren, die zur Familie des Nuklearfaktors-kappa-B (NF-kB) gehören, koordinieren die Expression wichtiger Gene, die an Immunantworten, Proliferation und Apoptose beteiligt sind. Zugleich deuten neue Erkenntnisse darauf hin, dass NF-kB bei der Entstehung und Progression von Krebs ebenfalls eine Rolle spielt. Um eine gegen NF-kB gerichtete Krebsbehandlung zu entwickeln, wäre es also nötig, selektiv auf diejenigen NF-kB-Gene abzuzielen, die mit maligner Transformation in Zusammenhang stehen. Vor diesem Hintergrund schlägt das EU-finanzierte Projekt RegulatioNFkB vor, den Mechanismus der durch NF-kB vermittelten transkriptionellen Aktivierung zu erforschen. Dazu wird es sich auf die Promotorbereiche von NF-kB-modulierten Genen konzentrieren. Mithilfe von Genomeditierung und Massenspektrometrie werden die Forschenden Transkriptionsfaktoren und weitere Proteine ermitteln, die an den Promotoren dieser Gene interagieren.

Ziel

NF-kB is a family of transcription factors that regulate expression of genes that are involved in various cellular processes such as apoptosis, proliferation and differentiation. The NF-kB pathway is implicated in the initiation and progression of cancer and cancer cell drug resistance. Unfortunately, global NF-kB inhibition is harmful to the immune system; therefore, inhibition of a subset of NF-kB target genes that are linked to malignant transformation would be more appropriate. Different NF-kB target genes display distinct transcription activation kinetics, indicating that the molecular mechanisms driving expression of NF-kB target genes are strikingly diverse. Thus, it is highly interesting to study the factors that are involved in transcriptional activation of individual NF-kB responsive genes, both from a fundamental and clinical perspective. Here, we will develop two innovative strategies based on CRISPR/Cas9 based genome targeting combined with mass spectrometry-based technology to isolate single NF-kB target gene promoters upon NF-kB pathway activation and identify their constituents. We will also apply a complementary approach, ATAC-sequencing, to identify transcription factors that are involved in activation of NF-kB responsive genes. Moreover, we will study the underlying mechanisms and characterize the function of proteins that interact with NF-kB responsive promoters. Finally, the described mechanisms discovered in cancer cell lines will be verified and validated in small intestinal organoids, a non-transformed cell model for homeostasis in the intestinal epithelium. This ambitious and multidisciplinary project will greatly benefit from my expert knowledge regarding the NF-kB pathway combined with the expertise of the host laboratory in mass spectrometry and genomics. The project is expected to have a major impact in the research area of gene expression regulation in general and molecular cancer research focusing on the NF-kB pathway in particular.