Hypothesis

Repeated sub‑lethal heat and acidification episodes induce a trained immunity state in coral host cells, characterized by stable H3K4me3 marks at promoters of innate immune genes. This epigenetic lock sustains TLR‑driven NF‑κB signaling after the stressor ends, driving chronic inflammaging, microbiome dysbiosis, and calcification decline. Blocking the histone methyltransferase responsible for this mark will reset immune homeostasis and extend holobiont healthspan.

Mechanistic Model

1. Stress‑induced signaling – Thermal spikes and low pH activate membrane TLRs and downstream MyD88‑dependent kinases, leading to transient ROS bursts and NF‑κB nuclear translocation.
2. Epigenetic imprinting – ROS‑sensitive signaling recruits the SET1/MLL complex (SETD1A homolog) to nucleosomes surrounding pro‑inflammatory gene promoters (e.g., tlr, ikk, caspase). This deposits H3K4me3, a mark associated with transcriptional readiness.
3. Persistent immune activation – Even after stressors subside, H3K4me3 maintains an open chromatin configuration, allowing rapid, amplified transcription upon minor stimuli. The result is a low‑grade, constitutive inflammatory state analogous to vertebrate inflammaging.
4. Holobiont feedback – Sustained NF‑κB drives expression of antimicrobial peptides and mucin modifiers that shift the surface microbiome toward opportunistic Vibrio spp., while suppressing beneficial Endozoicomonas. Dysbiotic microbes release PAMPs that further stimulate TLRs, creating a self‑reinforcing loop.
5. Phenotypic outcome – Chronic inflammation elevates matrix metalloprotease activity and inhibits carbonic anhydrase, reducing calcification rates and accelerating tissue loss, especially in older colonies with lower regenerative capacity.

Testable Predictions

• Corals exposed to a single heat‑acid pulse will show transient H3K4me3 enrichment at immune gene promoters, returning to baseline within 48 h.
• Corals subjected to repeated pulses (3× over 2 weeks) will retain elevated H3K4me3 marks for ≥10 days post‑stress, correlating with sustained tlr and nfkb transcription.
• Pharmacological inhibition of SETD1A activity (using a specific small‑molecule inhibitor) during the repeated‑pulse regimen will prevent H3K4me3 accumulation, normalize immune gene expression, maintain a Endozoicomonas-rich microbiome, and preserve calcification rates compared with untreated controls.
• Older (≥10 yr) Porites colonies will exhibit a higher baseline H3K4me3 load at immune loci than younger colonies, predicting greater susceptibility to inflammaging under identical stress regimes.

Experimental Approach

1. Stress regime – Maintain nubbins from juvenile and adult Porites colonies in flow‑through tanks. Apply either a single 3‑day 32 °C/pH 7.8 pulse or three pulses separated by 5‑day recovery periods.
2. Epigenetic assay – Perform ChIP‑qPCR for H3K4me3 at promoter regions of tlr2, ikkβ, and caspase3 using antibodies validated for cnidarian chromatin. Sample at 0 h, 24 h, 72 h, and 168 h after the final stressor.
3. Pharmacological intervention – Add SETD1A inhibitor (e.g., MM‑102 analog) to the water during the recovery intervals of the repeated‑pulse group; include vehicle controls.
4. Holobiont readouts – Quantify Endozoicomonas and Vibrio spp. via 16S rRNA amplicon sequencing; measure calcification buoyant weight; assess tissue health via live‑dead imaging.
5. Data analysis – Use mixed‑effects models to test interactions between age, stress frequency, inhibitor presence, and epigenetic/phenotypic outcomes. Falsification occurs if inhibitor treatment fails to reduce H3K4me3 accumulation or does not improve microbiome stability and calcification relative to controls.