Hypothesis

In aged adipose tissue, a stiffened extracellular matrix (ECM) activates integrin‑β1‑FAK signaling, which promotes nuclear translocation of YAP/TAZ. Once in the nucleus, YAP/TAZ recruit histone deacetylase 3 (HDAC3) to autophagy‑gene promoters (e.g., Map1lc3b, Becn1, Atg5), leading to chromatin condensation and transcriptional repression. This active suppression of autophagy sustains a pro‑fibrotic, senescent adipocyte state, whereas genetic or pharmacological disruption of the YAP/TAZ‑HDAC3 axis restores autophagic flux and ameliorates age‑associated adipose dysfunction.

Mechanistic Reasoning

1. ECM stiffness → integrin‑β1/FAK – Fibrotic collagen accumulation in aged adipose tissue increases tensile stress on adipocytes, engaging integrin‑β1 and focal adhesion kinase (FAK) {1}. FAK autophosphorylation (Y397) triggers downstream Src activation, which phosphorylates YAP on S127, inhibiting its cytoplasmic retention and favoring nuclear import {2}.
2. Nuclear YAP/TAZ as transcriptional repressors – While YAP/TAZ are known co‑activators of pro‑growth genes, recent work shows they can recruit corepressor complexes in a context‑dependent manner. In stiff microenvironments, YAP/TAZ interact with HDAC3 via its deacetylase domain, forming a repressor complex that binds TEAD motifs adjacent to autophagy promoters {3}.
3. HDAC3‑mediated chromatin silencing – HDAC3 removes acetyl groups from histone H3K27ac, promoting nucleosome compaction and reducing RNA polymerase II occupancy at autophagy loci. This transcriptional dampening lowers LC3‑II conversion and p62 degradation, measurable as reduced autophagic flux {4}.
4. Feedback loop – Suppressed autophagy leads to accumulation of damaged mitochondria and lipid droplets, which further stimulate ROS‑dependent TGF‑β secretion from macrophages, reinforcing ECM deposition and sustaining the mechanotransductive signal {5}.

Testable Predictions

• Prediction 1: In adipocytes isolated from old (≥24 mo) mice, nuclear YAP/TAZ levels will correlate positively with ECM stiffness (measured by atomic force microscopy) and inversely with LC3‑II/I ratio after bafilomycin A1 treatment.
• Prediction 2: Pharmacological inhibition of FAK (PF‑573228) or genetic adipocyte‑specific deletion of Itgb1 in aged mice will reduce nuclear YAP/TAZ, decrease HDAC3 recruitment to Map1lc3b promoter (ChIP‑qPCR), and restore autophagic flux without altering mTORC1 activity (p‑S6K unchanged).
• Prediction 3: Overexpression of a HDAC3‑resistant YAP mutant (YAP‑5SA) in young adipocytes will mimic the aged phenotype: increased nuclear YAP, HDAC3 recruitment, suppressed autophagy, and heightened expression of fibrotic markers (Col1a1, Acta2).
• Prediction 4: Treating aged adipocyte cultures with a selective HDAC3 inhibitor (RGFP966) will increase histone acetylation at autophagy promoters, elevate LC3‑II turnover, and reduce senescence‑associated β‑galactosidase activity, even in the presence of a stiffened matrix.

Experimental Approach (outline)

1. Biomechanics & signaling – Harvest epididymal fat from young (3 mo) and old (24 mo) mice; perform AFM to map elastic modulus; isolate stromal vascular fraction and adipocytes for Western blot of p‑FAK (Y397), total YAP/TAZ, nuclear/cytoplasmic fractionation.
2. Chromatin analysis – Conduct ChIP for YAP, TEAD1, and HDAC3 on Map1lc3b and Becn1 promoters; quantify H3K27ac levels.
3. Autophagy flux – Treat ex vivo adipocytes with bafilomycin A1 or chloroquine; measure LC3‑II and p62 by immunoblot; use mCherry‑GFP‑LC3 reporter to assess autophagosome‑lysosome fusion via fluorescence microscopy.
4. In vivo validation – Generate adipocyte‑specific Itgb1^fl/fl;Adipoq‑Cre mice; assess ECM histology (Masson’s trichrome), autophagy markers, and metabolic phenotypes (glucose tolerance, adipocyte size) at 24 mo.
5. Pharmacologic rescue – Treat aged mice with FAK inhibitor or HDAC3 inhibitor for 4 weeks; repeat autophagy and senescence readouts.

Falsifiability

If nuclear YAP/TAZ does not increase with ECM stiffness, or if HDAC3 inhibition fails to restore autophagy despite persistent stiff matrix, the hypothesis would be refuted. Conversely, demonstration that autophagy suppression persists when YAP/TAZ‑HDAC3 interaction is genetically disrupted would also falsify the proposed mechanism.

Broader Implications

This mechanism positions the adipocyte ECM as a rheostat that couples mechanical cues to epigenetic control of cellular housekeeping. It suggests that anti‑fibrotic strategies targeting integrin‑FAK‑YAP/TAZ‑HDAC3 signaling could rejuvenate adipose autophagy, mitigate inflammaging, and improve metabolic health in older individuals.