Signalling pathways required for Normal Fibroblast (NF) to Cancer-Associated Fibroblast (CAF) conversion


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Nil EGE1, Fernando Calvo1, Erik Sahai1
1Cancer Resarch UK, London, UK

Background

The microenvironment of the tumour or stroma is a complex network of different cell types. Cancer-associated fibroblasts (CAF) are one important population present in the stroma. As well as producing the extracellular matrix, they are key regulators of paracrine signalling between stromal and cancer cells. Several studies have investigated their origin, one hypothesis being that they arise from the conversion of resident normal fibroblasts (NF). However the molecular and functional differences between normal and cancer-associated fibroblasts have yet to be fully determined.

Method

Gene set enrichment analysis was completed to identify signaling pathways up-regulated in CAF compared to NF. Reporters to track transcriptional activities and protein localisations of these pathways are used for live cell imaging both in vitro and in vivo.

Results

Genomic analyses suggested that YAP, SRF and TGFbeta pathways are up regulated in CAF compared to NF. This was then confirmed by QPCR analysis studying the expression levels of target genes.
We have further identified several cytoskeletal regulators (such as ANLN, DIAPH3 or MYL9) specifically regulated by YAP and required for CAF functionality in organotypic cultures (Calvo et al.).
We are now testing the dynamic and specificity of our constructs to report transcriptional activities and protein localisation of YAP, SRF and TGFbeta pathways. We aim to use those reporters to visualise pathways activities in vitro and in vivo and to understand their chronology and their possible interdependence during fibroblast conversion.

Conclusion

In this project we have documented the role of YAP signaling pathway in CAF function. In parallel we set up several tools to further image in vitro and in vivo these three pathways activity during the activation and maintenance of CAF.