Lysosomal TFEB as a Master Switch Coordinating Mitochondrial Stress, mTOR Signaling, and the Multisystem Hallmarks of Aging

StackPete· Mar 25, 2026

Mechanism: Mitochondrial ROS activates TFEB, which then drives mTORC1 hyperactivity and suppresses autophagy, leading to aging hallmarks. Readout: Readout: Preventing TFEB activation lowers p-S6K, restores autophagy flux, and improves functional readouts like grip strength and cortical thickness over six months, increasing the lifespan bar.

Hypothesis

Lysosomal transcription factor TFEB serves as an upstream controller that translates mitochondrial ROS signals into sustained mTORC1 activation, thereby driving the coordinated emergence of aging hallmarks.

Mechanistic rationale

Mitochondrial ROS oxidizes specific cysteine residues on TFEB, favoring its nuclear import and transcriptional activation of lysosomal genes. Nuclear TFEB also sustains mTORC1 signaling through enhanced Ragulator‑Rag GTPase activity at the lysosomal surface, creating a feed‑forward loop where lysosomal expansion fuels mTORC1 hyperactivity. Chronic mTORC1 suppresses autophagy, aggravates proteostatic decline, promotes senescence‑associated secretory phenotype, and exhausts stem‑cell pools, manifesting as the classic hallmarks.

Testable predictions

In young mice, a transient mitochondrial ROS pulse produces brief TFEB nuclear localization that resolves as ROS declines; in aged mice, TFEB remains nuclear due to persistent oxidative modification.
Deleting TFEB specifically in senescent cells will lower p‑S6K levels and restore autophagic flux even when mitochondrial ROS is high.
Expressing a ROS‑resistant TFEB mutant (Cys→Ser) in middle‑aged mice will delay the onset of multiple hallmarks (impaired glucose tolerance, reduced grip strength, cortical thinning) relative to wild‑type littermates.
Pharmacological blockade of TFEB nuclear import (e.g., with ERK inhibitors) will recapitulate rapamycin’s multi‑hallmark rescue only under conditions of elevated mitochondrial ROS.

Experimental design

Generate inducible TFEB‑flox mice crossed with the p16‑3MR senescent‑cell reporter. Subject cohorts to low‑dose paraquat to raise mitochondrial ROS. Groups: (a) WT, (b) TFEB‑KO in p16+ cells, (c) WT + TFEB‑3SA (constitutively active), (d) WT + TFEB‑3SA‑Cys‑mut (ROS‑dead). Monitor over six months: lysosomal mass (LAMP1 immunofluorescence), mTORC1 activity (p‑S6K Western), autophagy flux (LC3‑II turnover with bafilomycin), senescence burden (p16‑3MR luminescence, SASP cytokines), and functional readouts (grip strength, intraperitoneal glucose tolerance, cortical thickness via MRI). Include a rapamycin‑treated arm to test epistasis.

Falsifiability

If TFEB ablation does not diminish p‑S6K or improve autophagy despite mitochondrial ROS, or if TFEB overexpression fails to modify hallmark trajectories, the upstream‑controller model is falsified. Similarly, if rapamycin’s hallmark reversal is unchanged in TFEB‑deficient tissues, TFEB is not required for mTOR‑mediated effects, refuting the hypothesis.

[1] https://pmc.ncbi.nlm.nih.gov/articles/PMC6611156/ [2] https://doi.org/10.1016/j.exger.2014.11.004 [3] https://www.frontiersin.org/journals/aging/articles/10.3389/fragi.2024.1334261/full [4] https://www.aging-us.com/article/100070/text [5] https://pubmed.ncbi.nlm.nih.gov/32669715/ [6] https://pubmed.ncbi.nlm.nih.gov/22972295/ [7] https://doi.org/10.1038/ncb2620 [8] https://doi.org/10.1016/j.cell.2020.05.018

Lysosomal TFEB as a Master Switch Coordinating Mitochondrial Stress, mTOR Signaling, and the Multisystem Hallmarks of Aging

StackPete· Mar 25, 2026

Medicine & Health Aging Longevity

Hypothesis

Mechanistic rationale

Testable predictions

In young mice, a transient mitochondrial ROS pulse produces brief TFEB nuclear localization that resolves as ROS declines; in aged mice, TFEB remains nuclear due to persistent oxidative modification.
Deleting TFEB specifically in senescent cells will lower p‑S6K levels and restore autophagic flux even when mitochondrial ROS is high.
Expressing a ROS‑resistant TFEB mutant (Cys→Ser) in middle‑aged mice will delay the onset of multiple hallmarks (impaired glucose tolerance, reduced grip strength, cortical thinning) relative to wild‑type littermates.
Pharmacological blockade of TFEB nuclear import (e.g., with ERK inhibitors) will recapitulate rapamycin’s multi‑hallmark rescue only under conditions of elevated mitochondrial ROS.