Receptor‑Specific Autophagy Triage Determines ECM Homeostasis and Is Disrupted in Aging

Hypothesis

Selective autophagy does not follow a universal cargo hierarchy; instead, parallel receptor pathways (p62/SQSTM1, NBR1, OPTN, SEC62) compete for ubiquitin chains within phase‑separated condensates, and the outcome of this competition dictates which substrate—ER, mitochondria, or ECM‑associated proteins—is degraded first. With advancing age, alterations in receptor phosphorylation and ubiquitin chain topology shift the balance toward preferential ER‑phagy, suppressing mitophagy and ECM‑protective autophagy, leading to accumulated mitochondrial damage and maladaptive matrix remodeling.

Mechanistic Reasoning

• p62 and NBR1 form cooperative condensates that bind ubiquitinated cargo; NBR1 acts as a chain terminator, promoting condensate growth, while p62 provides scaffolding for ULK1 recruitment (https://pmc.ncbi.nlm.nih.gov/articles/PMC12669772/, https://pmc.ncbi.nlm.nih.gov/articles/PMC12740005/).
• In inflammatory muscle atrophy, OPTN and SEC62 are up‑regulated early, indicating ER‑phagy precedes mitochondrial turnover (https://doi.org/10.1101/2025.05.06.650963).
• Parkin‑dependent mitophagy accounts for only ~25% of mitochondrial protein turnover; BNIP3/NIX‑mediated pathways dominate under hypoxia (https://doi.org/10.1080/15548627.2019.1586258).
• FGF21‑driven autophagy improves ECM homeostasis via SIRT1‑mTOR signaling, not direct collagen degradation (https://doi.org/10.1038/s41419-021-04157-x).

We propose that aging modifies the ubiquitin landscape (e.g., increased K63‑linked chains on ER‑resident proteins) and alters kinase activity (e.g., heightened TBK1/IKKε phosphorylation of OPTN and p62). These changes bias condensate composition toward OPTN/SEC62‑rich assemblies, favoring ER sequestration. Consequently, mitophagic receptors (BNIP3/NIX, Parkin) are outcompeted for limited autophagosome initiation sites, and FGF21‑stimulated autophagy flux is diverted away from ECM‑protective substrates.

Testable Predictions

1. Condensate Profiling – In young vs. aged murine lung fibroblasts, immuno‑EM will show a higher ratio of OPTN/SEC62 to p62/NBR1 within ubiquitin condensates, correlating with increased ER‑phagy markers (FAM134B‑LC3 colocalization) and decreased mitochondrial (TOM20‑LC3) and collagen‑II‑LC3 signals.
2. Phospho‑Swap Rescue – Expressing a phosphorylation‑deficient OPTN mutant (S177A) in aged cells will restore p62/NBR1 dominance, normalize mitophagy (mt‑Keima assay), and reduce MMP13/ADAMTS5 expression (qPCR, gelatin zymography).
3. Ubiquitin Chain Bias – Linkage‑specific ubiquitin antibodies will reveal enriched K63 chains on ER‑shaping proteins (RTN4) and diminished K48 chains on mitochondrial outer‑membrane proteins (MFN2) in aged tissues; treating cells with the K63‑specific deubiquitinase inhibitor NSC632839 will shift cargo preference back to mitochondria.
4. Functional Readout – Aged mice treated with an OPTN/SEC62‑targeting siRNA (delivered via lung‑specific nanoparticles) will exhibit improved collagen II deposition (Sir‑Red staining) and enhanced respiratory compliance (flexiVent), without altering global LC3‑II levels.

If these experiments fail to show a shift in receptor‑biased cargo selection or rescuing the phosphorylation state does not mitigate ECM deterioration, the hypothesis would be falsified, indicating that age‑related decline stems from global autophagy insufficiency rather than altered triage hierarchy.