Phosphoproteomics of cellular mechanosensing reveals NFATC4 as a regulator of myofibroblast activity

ABSTRACT: Feedback connections between tissue stiffness and cellular contractile forces can instruct cell identity and activity via a process referred to as mechanosensing. Specific phosphoproteome changes during mechanosensing are poorly characterized. In this work, we chart the global phosphoproteome dynamics of primary human lung fibroblasts sensing the stiffness of injury relevant fibronectin coated Poly(dimethylsiloxane) substrates. We discovered a key signaling threshold at a Young's modulus of eight kPa stiffness, above which cells activated a large number of pathways including RhoA, CK2A1, PKA, AMPK, AKT1, and Hippo-YAP1/TAZ mediated signaling. Time-resolved phosphoproteomics of cell spreading on stiff substrates revealed the temporal dynamics of these stiffness-sensitive signaling pathways. ECM substrate stiffness above eight kPA induced fibroblast contractility, cytoskeletal rearrangements, ECM secretion, and a fibroblast to myofibroblast transition. Our data indicate that phosphorylation of the transcriptional regulator NFATC4 at S213/S217 enhances myofibroblast activity, which is the key hallmark of fibrotic diseases. NFATC4 knock down cells display reduced stiffness induced collagen secretion, collagen gel contraction, and cell invasion, suggesting NFATC4 as a novel target for antifibrotic therapy.

INSTRUMENT(S):

ORGANISM(S): Homo Sapiens (human)

TISSUE(S): Cell Culture, Fibroblast

DISEASE(S): Idiopathic Pulmonary Fibrosis

SUBMITTER: Safouane Kadri  

LAB HEAD: Herbert Schiller

PROVIDER: PXD063448 | Pride | 2026-06-02

REPOSITORIES: Pride