The Hippo pathway links adipocyte plasticity to adipose tissue fibrosis

Adipose tissue fibrosis, characterized by excessive ECM deposition, accompanies maladaptive remodeling during obesity and contributes to insulin resistance and metabolic disease. While adipocytes lose aspects of fat identity under nutrient excess and some adopt ECM-remodeling features, the mechanisms connecting adipocyte plasticity to fibrosis remain unclear. TGFβ is a central driver of fibrosis via canonical SMAD2/3 and non-canonical pathways, and the Hippo pathway controls tissue homeostasis by restraining YAP/TAZ activity. Prior work implicates YAP/TAZ in adipogenesis decisions, but whether Hippo signaling governs adipocyte plasticity and fibrosis in vivo was unknown. This study tests the hypothesis that Hippo pathway inactivation in adipocytes, in concert with TGFβ signaling, drives adipocyte dedifferentiation toward profibrotic states and promotes adipose tissue fibrosis, with implications for obesity-associated dysfunction.

• Adipose tissue remodeling during obesity includes hypertrophy, inflammation, and fibrosis and is linked to metabolic dysfunction. Prior studies show adipocyte hypertrophy reduces fat-identity gene expression and increases inflammatory adipokines. Fibrosis correlates with insulin resistance in obesity.
• Known regulators of AT fibrosis include hypoxia/HIF1α (induces ECM genes and collagen crosslinking enzymes like LOX), PRDM16 complexes (limit fibrosis under adrenergic stimulation and signal to precursors to block myofibroblast differentiation), and PDGFRα signaling (drives fibrosis via perivascular cells and PDGFRα+CD9high progenitors). A subpopulation of mature adipocytes can shift to ECM-remodeling states during HFD.
• TGFβ (three isoforms) canonically signals via TGFBR2/TGFBR1 to phosphorylate SMAD2/3, inducing ECM genes; non-canonical MAPK, PI3K/AKT, and Rho/ROCK pathways also contribute to fibrosis.
• The Hippo pathway restricts YAP/TAZ; when LATS1/2 activity is low, YAP/TAZ enter the nucleus to coactivate TEADs. YAP/TAZ influence mesenchymal lineage choice (adipogenesis vs osteogenesis) and have roles in adipocyte differentiation in vitro, but their in vivo role in adipocyte plasticity and fibrosis had been unclear.

• Mouse genetics: Generated adipocyte-specific Lats1/2 double knockout (L1L2-AKO; Lats1fl/fl;Lats2fl/fl;Adipoq-Cre) and inducible adipocyte-specific knockout (L1L2-iAKO; Adipoq-CreERT2). Created Yap/Taz adipocyte knockout (YT-iAKO; Yap1fl/fl;Tazfl/fl;Adipoq-CreERT2) and compound Lats1/2/Yap/Taz knockouts (L1L2YT-AKO, and single Yap- or Taz-rescue variants). Also generated Mst1/2 adipocyte knockout (M1M2-AKO) to test upstream Hippo kinases.
• Obesity models: High-fat diet (HFD) feeding for up to 18 weeks; genetic ob/ob mice.
• AAV tools and gene delivery: Local scWAT injections of AAV vectors driven by adiponectin promoter (ADP) for adipocyte targeting. Overexpressed active TAZ(4SA) or YAP(5SA). Used AAV-ADP-Cre to delete Lats1/2 in adult adipocytes. Activated TGFβ signaling via constitutively active TGFBR1(T204D) using CAG, DIO (Cre-dependent), or FIO (FLPo-dependent) constructs to achieve adipocyte-restricted expression in L1L2-iAKO mice. Overexpressed active TGFβ1(2CS). Delivered CCL2/CCL7 to recruit macrophages. Verified LATS1/2 knockdown by immunoblot.
• Cell culture and CRISPR: Differentiated SVF-derived adipocytes from scWAT; CRISPR/Cas9 AAV gRNAs to knockout Lats1/2 and/or Yap/Taz or Tead1-4 in vitro; assessed adipocyte identity and fibrosis markers by RT-qPCR and immunoblot.
• Histology and imaging: H&E for adipocyte morphology; Masson’s trichrome and Picrosirius red for collagen deposition; immunofluorescence for αSMA, p-SMAD2, COL1A1, DPP4; subcellular fractionation for YAP/TAZ localization.
• Flow cytometry: SVF profiling of DPP4+ and DPP4− progenitors; macrophage populations (F4/80, CD11c, CD206); live gating on CD45−CD31− for stromal cells. Magnetic sorting of F4/80+ macrophages for Tgfb1/2/3 expression analysis.
• Fate mapping and transplantation: Used LSL-Cas9-EGFP or mT/mG reporters in L1L2-AKO to trace adipocyte-derived cells; sorted GFP+DPP4+/- populations; in vitro TGFβ1 treatment to test conversion to myofibroblasts (αSMA+); transplanted GFP+DPP4+ cells into recipient fat pads and tracked fate.
• Signaling assays: Western blot for Hippo (LATS1/2, YAP/TAZ, p-YAP/TAZ) and TGFβ (SMAD2/3 and p-SMAD2/3) components; cycloheximide chase to assess SMAD2 protein stability; ubiquitination IP assays for SMAD2; assessment of MAPK/AKT/ROCK pathways.
• Human data: Transcriptome analysis of human scWAT for fibrosis and YAP signature; correlation of WWTR1/YAP1 with COL6A3/MMP2.
• Pharmacology and metabolic tests: Verteporfin (25 mg/kg) to disrupt YAP/TAZ-TEADs; glucose and insulin tolerance tests; DXA for body composition; serum NEFA quantification.
• Statistics: Student’s t-test, one-/two-way ANOVA with Bonferroni correction; data as mean ± SEM.

• Obesity associates with Hippo inactivation and fibrosis in adipose tissue: In ob/ob and HFD-fed mice, fibrotic genes (e.g., Col1a1, Col6a1, Mmp2) and YAP/TAZ targets (Ctgf, Cyr61) are elevated. LATS2 protein is reduced, YAP/TAZ increased, and p-YAP/p-TAZ ratios decrease, indicating pathway inhibition. Human scWAT shows similar upregulation of fibrosis and conserved YAP signatures; WWTR1/YAP1 expression correlates with COL6A3/MMP2.
• Adipocyte-specific Lats1/2 deletion (L1L2-AKO) causes severe fat remodeling: Marked fat mass loss without apoptosis, reduced adipocyte identity genes (Pparg, Fabp4, Adipoq), reduced HSL/Perilipin1, and strong induction of fibrotic programs (Col1a1, Col3a1, Mmp2, Mmp14, Fn1, Lox, Tagln, Acta2/αSMA) with extensive collagen deposition by 5 weeks. Single Lats1 or Lats2 knockout does not recapitulate phenotype, indicating redundancy.
• Lats1/2 deficiency-induced fibrosis is MST1/2-independent: Adipocyte-specific Mst1/2 deletion does not induce fat loss or fibrosis.
• Hippo inactivation alone in adult adipocytes is insufficient for fibrosis: Inducible L1L2-iAKO or adipocyte YAP(5SA)/TAZ(4SA) overexpression in adults impair adipocyte identity and cause fat loss but do not induce robust fibrosis.
• TGFβ cooperation required: TGFβ pathway is more active in early postnatal AT; L1L2-AKO shows elevated Tgfb and p-SMAD2. In adults, activating TGFβ signaling (TGFBR1 T204D or TGFβ1 2CS) together with Lats1/2 deletion triggers strong fibrotic responses (αSMA+ myofibroblasts, ECM genes). TGFβ activation alone in controls increases p-SMAD2 but is insufficient to cause fibrosis.
• Obesity provides endogenous TGFβ signal that synergizes with Hippo inactivation: In obese settings (HFD or ob/ob), adipocyte-targeted Lats1/2 deletion via AAV-ADP-Cre further induces fibrosis markers and collagen deposition compared to controls.
• Adipocyte-to-myofibroblast lineage transition: In L1L2-AKO, a subset of αSMA+ myofibroblasts derive from adipocytes that dedifferentiate into DPP4+ progenitors. GFP+ adipocyte-derived cells become DPP4+ early and later αSMA+; in vitro, Lats1/2-deficient adipocytes upregulate Dpp4; TGFβ converts GFP+ DPP4− cells to DPP4+ αSMA+ cells.
• Immune-stromal crosstalk: Lats1/2 deficiency elevates Ccl2/Ccl7 and recruits macrophages (F4/80+), which produce TGFβ, establishing a CCL2/7–macrophage feedforward loop that amplifies fibrosis. Overexpressing CCL2/CCL7 increases macrophage markers and fibrosis genes in L1L2-iAKO.
• Mechanism: YAP/TAZ-TEADs are necessary for adipocyte identity loss and fibrosis. Deleting Yap/Taz rescues adipocyte identity and abrogates fibrosis in L1L2-AKO. TEAD1-4 knockout rescues identity and prevents YAP/TAZ nuclear accumulation. Hippo inactivation increases SMAD2 protein (not mRNA) via YAP/TAZ-dependent stabilization by reducing SMAD2 ubiquitination, prolonging its half-life; loss of Yap/Taz increases SMAD2 ubiquitination.
• Therapeutic targeting: Inducible adipocyte-specific Yap/Taz deletion after fibrosis onset reduces ECM accumulation and improves glucose tolerance/insulin sensitivity in HFD mice. Verteporfin treatment in ob/ob and HFD models reduces fibrotic gene expression, collagen deposition, YAP targets, and improves metabolic tests. Conversely, adipocyte TAZ(4SA) overexpression exacerbates fibrosis in obesity.

The study demonstrates that Hippo pathway inactivation in adipocytes links cellular plasticity to adipose tissue fibrosis through synergy with TGFβ signaling. In obesity, partial Hippo inactivation (reduced LATS activity, increased YAP/TAZ) and elevated TGFβ signaling converge to impair adipocyte identity and promote ECM remodeling. Complete adipocyte Lats1/2 loss in postnatal mice induces dedifferentiation into DPP4+ progenitors that, upon TGFβ stimulation and macrophage-derived signals, transition to myofibroblasts. Mechanistically, YAP/TAZ activation stabilizes SMAD2 protein by limiting its ubiquitination, sensitizing cells to TGFβ and sustaining fibrotic gene transcription via YAP/TAZ-TEAD–SMAD2 cooperation. The identification of a CCL2/CCL7–macrophage loop that augments TGFβ further explains how inflammation accelerates fibrosis. These findings address the open question of how adipocyte plasticity contributes to fibrosis and highlight Hippo–TGFβ integration as a central rheostat controlling adipose remodeling, with broad relevance to metabolic disease.

This work establishes the Hippo pathway as a molecular switch governing adipocyte identity and fate, and reveals a two-signal model in which Hippo inactivation cooperates with TGFβ to drive adipocyte dedifferentiation and myofibroblast conversion, promoting adipose tissue fibrosis. YAP/TAZ-TEADs stabilize SMAD2 to potentiate TGFβ responses, while macrophage recruitment via CCL2/CCL7 amplifies fibrotic signaling. Therapeutically, genetic inactivation of YAP/TAZ in adipocytes or pharmacological disruption of YAP/TAZ–TEADs with verteporfin alleviates obesity-induced fibrosis and improves metabolic homeostasis. Future studies should delineate physiological upstream regulators of LATS1/2 (e.g., MST1/2, MAP4Ks, mechanical cues, lipids) in adipose tissue and assess translatability to human adipose fibrosis.

Adipocyte-specific Lats1/2 deletion induces severe lipodystrophy not typically observed physiologically, potentially limiting direct extrapolation. The model may overstate the extent of Hippo pathway inactivation compared to obesity. While MST1/2 were not required here, other upstream regulators (e.g., MAP4Ks, mechanical stress, free fatty acids) likely modulate Hippo activity in vivo and were not fully dissected. The precise contribution of adipocyte-derived versus precursor-derived myofibroblasts needs further quantification. Pharmacological targeting with verteporfin, though effective, is not YAP/TAZ-specific and off-target effects cannot be excluded.