Hypothesis

KDM6A functions as a direct chromatin effector of nutrient status, demethylating H3K27me3 at bivalent promoters to enable a transcriptional program that suppresses mTORC1 and activates survival pathways. When nutrients are abundant, KDM6A activity is low, H3K27me3 remains high at loci such as Deptor, mTORC1 stays active, and cells pursue growth and specialization (the 'civilization' state). Under nutrient restriction or stress, AMPK-activated KDM6A increases H3K27me3 removal, facilitating Deptor expression, mTORC1 inhibition, and a shift toward autophagy, stress resistance, and maintenance (the 'survival' state). Loss of KDM6A locks cells in a constitutive mTORC1‑high, chromatin‑rigid configuration that mimics perpetual civilization signaling, thereby accelerating aging phenotypes and tumorigenesis.

Mechanistic Model

1. Nutrient sensing – Low glucose/amino acids activate AMPK, which phosphorylates KDM6A (predicted site Sxxx) enhancing its nuclear localization and demethylase activity.
2. Chromatin remodeling – Activated KDM6A binds bivalent promoters marked by both H3K27me3 and H3K4me3, removing H3K27me3 while preserving H3K4me3, creating a permissive state for transcription.
3. Deptor upregulation – The Deptor promoter loses H3K27me3, gains RNA Pol II occupancy, and its mRNA rises, leading to Deptor protein that binds and inhibits mTORC1 complexes.
4. mTORC1 suppression – Reduced mTORC1 signaling decreases p‑S6K and p‑4EBP1, relieving inhibition of ULK1 and TFEB, thereby stimulating autophagy and lysosomal biogenesis.
5. Feedback – Autophagic flux replenishes metabolites, further modulating AMPK activity and KDM6A phosphorylation, completing a nutrient‑chromatin‑metabolism loop.

Testable Predictions

• In wild‑type cells, acute amino acid starvation will increase KDM6A chromatin occupancy at the Deptor promoter and other bivalent loci, concomitant with H3K27me3 loss and H3K4me3 retention.
• KDM6A knockout or phospho‑deficient mutants will fail to show these chromatin changes under starvation, maintaining high H3K27me3, low Deptor expression, and sustained mTORC1 activity.
• Restoring Deptor expression in KDM6A‑deficient cells will rescue mTORC1 inhibition and autophagy induction despite the chromatin defect.
• Chronic mTORC1 inhibition (e.g., rapamycin) will not further decrease H3K27me3 at KDM6A‑target promoters in KDM6A‑null cells, indicating that KDM6A acts upstream of mTORC1 in this axis.

Experimental Design

1. Cell models – Primary mouse hepatocytes and human fibroblasts engineered with CRISPR‑Cas9 KDM6A knockout, phospho‑dead (SxxxA) and phospho‑mimetic (SxxxD) knock‑in lines.
2. Nutrient manipulations – Serum‑free, low‑glucose, or amino‑acid‑free media for 2‑6 h; parallel rapamycin (100 nM) treatment as pharmacologic mTORC1 inhibition control.
3. Assays –
   • ChIP‑seq for H3K27me3, H3K4me3, and KDM6A.
   • RNA‑seq (focus on Deptor and mTORC1‑responsive genes).
   • Western blot for p‑S6K, p‑4EBP1, LC3‑II/I, p62, and nuclear TFEB.
   • Autophagy flux assay (mCherry‑GFP‑LC3).
   • Senescence markers (SA‑β‑gal, p16^INK4a^).
4. Epistasis – Overexpress Deptor via lentiviral vector in KDM6A‑null cells; assess whether mTORC1 suppression and autophagy are restored.

Potential Outcomes and Implications

If the hypothesis holds, we will demonstrate that KDM6A translates nutrient status into a chromatin state that directly gates the mTORC1‑dependent civilization‑versus‑survival decision. This would position KDM6A as a lynchpin linking epigenetics, metabolism, and aging, offering a novel target for interventions that promote survival programs without the broad immunosuppression associated with chronic mTOR inhibition. Conversely, failure to observe nutrient‑dependent KDM6A chromatin dynamics would refute the model and redirect focus toward alternative mediators (e.g., KDM6B/HIF‑1) in the mTOR‑chromatin axis.