Epigenetic analysis of murine normal mammary gland fibroblasts (mNFs) and MMTV-PyMT cancer-associated fibroblasts (mCAF1) (H3K27ac ChIP-seq)

In comparison with normal tissue fibroblast (normal fibroblasts, NFs), cancer-associated fibroblasts (CAFs) have pathologically activated phenotypes that impact in a wide variety of tumoral processes including cancer cell growth and invasion, extracellular matrix (ECM) remodelling, angiogenesis and immune suppression. These phenotypes result from the establishment of altered gene expression programs that may be further sustained through epigenetic alterations. However, how epigenetic rearrangements influence CAF behaviour is not well delineated. Here, we identify aspartoacylase (ASPA) as a metabolic enzyme consistently repressed in tumour stroma and CAFs. We report a reciprocal crosstalk between ASPA and Transforming Growth Factor Beta (TGFβ) signalling that influences fibroblast behaviour. TGFβ suppresses ASPA expression in fibroblasts, whereas ASPA restrains TGFβ-dependent myofibroblast conversion, ECM remodelling, angiogenesis and pro-tumoral macrophage phenotypes. TGFβ/SMAD3-dependent transcriptional repression may require the activity of histone deacetylases (HDACs), which participate in epigenetic rearrangements modulating gene expression. Epigenomic analysis of the 3’ regulatory regions at the Aspa locus in murine MMTV-PyMT-derived breast cancer CAFs (mCAF1) as compared to normal murine mammary gland fibroblasts (mNFs), revealed a selective loss of H3K27Ac, an epigenetic mark of active enhancers. As expected, bona fide CAF marker genes such as Acta2, Fap, Lrrc15 and others presented increased H3K27Ac signal at regulatory regions in mCAF1, whereas peaks at NF marker genes (Cxcl1 and Pi16) were decreased. Our findings unveil ASPA expression in fibroblasts as a previously unknown gatekeeper of TGFβ responses and activation in cancer progression.