Hypothesis

Amyloid-like aggregates are not inert waste but a regulated, inducible reservoir of cryptic bioactive peptides that can be liberated by controlled proteolytic trimming to modulate stress‑responsive signaling pathways. When proteostasis capacity declines, the reservoir becomes overly stable, peptide release is impaired, and the loss of these signaling molecules contributes to functional decline.

Mechanistic Basis

• Aggregation‑prone proteins often contain disordered regions enriched in motifs that, upon proteolytic cleavage, generate peptides with hormetic activity (e.g., mitochondrial‑derived peptides, insulin‑like peptides).
• Sequestration into amyloid‑like fibrils protects the cell from toxic soluble oligomers while keeping these regions intact but inaccessible.
• Certain lysosomal proteases (cathepsins) or caspase‑like activities can partially unpack fibrils, releasing peptides that act extracellularly or intracellularly to activate stress‑resistance pathways such as HIF‑1α, NRF2, or FOXO.
• This process is reversible: during rejuvenation (e.g., partial OSKM reprogramming or lysosomal activation), increased protease activity and aggregate disaggregation liberate the peptide pool, resetting signaling without erasing the underlying protein sequences.

Testable Predictions

1. Inhibiting lysosomal protease activity in aged cells will increase aggregate stability and decrease the abundance of specific cryptic peptides derived from aggregated proteins, correlating with reduced stress resistance.
2. Exogenous addition of peptides liberated from purified age‑associated aggregates will recapitulate the protective effects of aggregate dissolution (e.g., enhanced survival under oxidative stress) even when aggregates remain intact.
3. Genetic manipulation to increase the proteolytic accessibility of aggregation‑prone regions (e.g., inserting protease‑sensitive linkers) will extend lifespan in model organisms by promoting peptide release without reducing overall aggregate load.

Experimental Approach

• Step 1: Generate C. elegans strains expressing a fluorescently tagged, aggregation‑prone protein (e.g., polyQ::GFP) alongside a protease‑sensitive reporter peptide fused to its C‑terminus.
• Step 2: Treat young and adult worms with lysosomal activators (e.g., TFEB overexpression) or inhibitors (e.g., chloroquine) and measure:
  • Aggregate load by fluorescence microscopy.
  • Released peptide levels by targeted mass‑spectrometry peptidomics.
  • Stress resistance assays (heat shock, oxidative stress).
• Step 3: Synthetically produce the candidate peptide identified in step 2 and add it to inhibitor‑treated worms; assess whether stress resistance is rescued despite persistent aggregates.
• Step 4: Perform epistasis analysis with insulin/IGF‑1 pathway mutants (daf‑2) to test whether peptide signaling acts downstream or parallel to known longevity regulators.

Potential Implications

If aggregates function as a peptide reservoir, therapeutic strategies that aim solely to dissolve aggregates may inadvertently deplete a beneficial signaling pool. Instead, modulating the kinetics of peptide release—enabling controlled liberation while maintaining aggregate sequestration—could preserve proteostatic order and extend healthspan. This reframes the anti‑aggregation paradigm from removal to regulated "tuning" of the amyloid‑based depot.

References

• [1] https://pmc.ncbi.nlm.nih.gov/articles/PMC7973223/
• [2] https://pmc.ncbi.nlm.nih.gov/articles/PMC6415910/
• [3] https://doi.org/10.1126/science.aag3048
• [4] https://journals.plos.org/plosbiology/article?id=10.1371%2Fjournal.pbio.1000450
• [5] https://elifesciences.org/articles/43059