PLPP4+ CAFs drive cancer progression by promoting ECM deposition via ceramide-mediated H3K27ac modification and CREB3L1 activation

Background Cancer-associated fibroblasts (CAFs) are the primary cellular source of extracellular matrix (ECM) in the tumor microenvironment (TME). Nevertheless, heterogeneity among CAF subsets results in substantial differences in their ECM-producing capacity, and definitive functional markers for the high ECM-producing CAF subset remain to be identified. Methods Integrated single-cell RNA sequencing (scRNA-seq) analysis stratified CAF subsets and identified subset-specific membrane markers, whose functions were validated via gain- and loss-of-function assays. Immunohistochemistry, immunofluorescence, Masson’s trichrome and Sirius Red staining characterized target protein localization and ECM accumulation. In vitro spheroid outgrowth and patient-derived organoid (PDO)-CAF co-culture assays, alongside in vivo syngeneic, xenograft and intrasplenic liver metastasis mouse models, assessed CAF-driven tumor growth and metastasis. Untargeted lipidomics, ChIP-qPCR, luciferase reporter and co-immunoprecipitation assays uncovered the underlying molecular mechanisms. Public scRNA-seq, spatial transcriptomic and clinical databases validated our findings across pan-cancer and fibrotic tissues. Results We identified PLPP4 as a functional marker of an ECM-enriched CAF subset. Functionally, PLPP4 drives tumor growth and metastasis both in vitro and in vivo by promoting excessive deposition of ECM. Mechanistically, PLPP4 generates ceramide via its phospholipid phosphatase activity, which upregulates ECM expression through a dual coordinated mechanism. On one hand, ceramide facilitates AMPK-mediated phosphorylation and subsequent ACSS2 nuclear import, thereby expanding the nuclear pool of acetyl coenzyme A (acetyl-CoA) and enhancing H3K27ac modification at the promoter regions of ECM-related genes. On the other hand, ceramide promotes the activation of CREB3L1, a transcription factor that governs the expression of multiple ECM genes. Together, these two pathways drive the excessive expression of ECM genes, including COL1A1 , COL1A2 , and fibronectin-1 ( FN1 ). Additionally, PLPP4 expression is upregulated in CAFs upon glutamine deprivation in an ATF4-dependent manner. Clinically, high PLPP4 expression in colorectal cancer correlates with denser ECM deposition, impaired immune cell infiltration, and unfavorable patient prognosis. Conclusions Collectively, our findings establish PLPP4⁺ CAFs as a critical tumor-promoting subset that drives cancer progression via enhanced ECM deposition, highlighting PLPP4 as a promising therapeutic target for anti-cancer strategies.