Critical role for the TGF-β1/mTORC1 signalling axis in defining the transcriptional identity of CTHRC1+ pathologic fibroblasts in IPF

<bold>Background:</bold> Fibroblasts represent key effector cells responsible for fibrogenesis during wound healing and pathological fibrosis, including idiopathic pulmonary fibrosis (IPF), the most rapidly progressive and fatal fibrotic lung disease. scRNA sequencing of fibrotic lung tissue has identified a novel population of high collagen-producing fibroblasts not present in control lung, characterised by high expression of the marker CTHRC1. The cardinal pro-fibrotic mediator TGF-β1 has been widely implicated in promoting fibrogenesis in multiple fibrotic conditions. In addition to the canonical Smad signalling pathway, we demonstrated that TGF-β1-induced collagen I production is under critical regulatory control by the mTORC1/4E-BP1 signalling hub. <bold>Methods & Results:</bold> Using pharmacological inhibition in combination with gene-editing approaches, we now demonstrate that the role of mTORC1 extends to the regulation of up to a third of all TGF-β1 regulated matrisome genes. We further show that the global transcriptome of TGF-β1-stimulated fibroblasts matches that of CTHRC1+ pathological fibroblasts in the IPF lung. In contrast, the TGF-β1 induced transcriptome of fibroblasts in which mTORC1-signalling is disrupted (by CRISPR-Cas9 <italic>RPTOR</italic> gene editing) does not map to any known fibroblast population. <bold>Conclusions:</bold> These data define, for the first time, a critical role for the TGF-β1/mTORC1 signalling hub in determining the transcriptional identity of CTHRC1+ pathological fibroblasts and provide strong scientific support for targeting mTORC1 as a therapeutic strategy in IPF and potentially other fibrotic conditions associated with dysregulated TGF-β1 signalling.