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

Descrizione del progetto

Analizzare il meccanismo dell’attivazione trascrizionale mediata dall’NF-kB

I fattori di trascrizione che rientrano nella famiglia del fattore nucleare kappa B (NF-kB, nuclear factor-kB) regolano l’espressione dei principali geni coinvolti nelle risposte immunitarie, nella proliferazione e nell’apoptosi. Al contempo, prove emergenti indicano che l’NF-kB riveste un ruolo attivo nell’insorgenza e nella progressione del cancro. Per sviluppare un trattamento antitumorale anti-NF-kB, pertanto, sarebbe necessario bersagliare selettivamente i geni dell’NF-kB associati alla trasformazione maligna. Alla luce di tali premesse, il progetto RegulatioNFkB, finanziato dall’UE, propone di approfondire il meccanismo alla base dell’attivazione trascrizionale mediata dall’NF-kB concentrando l’attenzione sulle regioni che promuovono i geni reattivi all’NF-kB. I ricercatori si avvarranno dell’editing genomico e della spettrometria di massa allo scopo di individuare i fattori di trascrizione e altre proteine che interagiscono nelle regioni promotrici di questi geni.

Obiettivo

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.