Per-species master-gene γ: an honest negative on grounding the cross-species loop gain

The osmoregulatory master gene ATP1A1 had its promoter γ measured across six species. The measurement is sound, but γ does not order species by loop gain — it tracks promoter GC, a lineage property — so the within-genome γ-ladder does not transfer across genomes. An honest negative: the comparative absolute gain stays open for a demonstrated reason.

The comparative chapter (§10) ordered animals by a single loop gain but left the absolute gains open and the per-species master-gene γ unmeasured. This chapter measures it — and reports an HONEST NEGATIVE. The osmoregulatory master gene ATP1A1 (the Na⁺,K⁺-ATPase α-1, the universal pump powering all secondary ion transport) had its promoter γ measured across six species spanning the osmotic-strategy axis, by the same NCBI / SantaLucia-1998 pipeline the rest of the framework uses. The measurement is sound, but γ does NOT order the species by their loop gain. It is published here as a negative, not hidden.

Measuring the osmoregulatory master gene across the strategy axis

From osmoconformers (Pacific oyster) through urea-retaining elasmobranchs (elephant shark, thorny skate) and aquatic regulators (zebrafish, Xenopus) to a terrestrial regulator (human), the ATP1A1 proximal-promoter γ = −mean nearest-neighbor stacking energy was measured from the public genome of each species — never fitted. Two internal checks confirm the pipeline is sound: the human ATP1A1 γ reproduces its standalone anchor (True), and the two independent elasmobranchs land within 0.0003 of each other (True) — a near-identical replicate.

The grounding test fails

For γ to ground the comparative loop gain k, it would have to increase with k across the strategy axis. It does not. The rank correlation is only Spearman(γ, k) = 0.647, and the ordering is plainly non-monotone: the two k=3.0 aquatic regulators alone span most of the entire conformer-to-mammal γ range, and the k=3.0 amphibian γ (1.4973) actually EXCEEDS the higher-gain k=4.0 mammal (1.4867). Per-species ATP1A1 γ does not grade the osmoregulatory loop gain.

Why: γ here is a GC readout, a lineage property

The diagnosis is clean. Across these six species γ tracks promoter GC content almost perfectly — Spearman(γ, GC) = 1.000. Nearest-neighbor stacking energy is dominated by GC pairs, so a same-gene promoter γ measured ACROSS genomes mostly reads each genome's background GC, a lineage / genome-composition property, not that species' osmoregulatory precision. The within-genome γ-ladder that works inside one organism (different master genes sharing a common GC background) does not transfer to a one-gene, many-genome comparison.

What this means

The comparative absolute loop gain k therefore STAYS open — but it is now open for a stated, demonstrated reason rather than as a bare placeholder: per-species master-gene γ is the wrong instrument for it, confounded by promoter GC. The measurement itself is verified [V] (NCBI, SantaLucia 1998, offline-reproducible, human-anchored), the negative result and the GC confound are verified [V], the strategy-to-clade assignment is cited [L], and the comparative absolute k is [O] with this confound as its documented obstacle. A genuine cross-species grounding would need a within-lineage or GC-controlled comparison — named here as the next instrument, not papered over.