rs5749131 — TCN2

Upstream regulatory variant near TCN2 associated with reduced holotranscobalamin levels — the bioactive fraction of circulating vitamin B12 available for cellular uptake

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TCN2 Upstream Variant — A Regulatory Dial on Your B12 Transport

Most vitamin B12 circulating in your blood is metabolically inert — bound to haptocorrin and unable to enter cells. Only 20–25% binds to transcobalamin II11 transcobalamin II
The only B12 carrier protein that delivers cobalamin into cells via the CD320 receptor on cell surfaces; encoded by the TCN2 gene on chromosome 22, forming holotranscobalamin22 holotranscobalamin
Also called "active B12" or holoTC — the fraction of circulating B12 that is actually deliverable to tissues. HoloTC below 35–50 pmol/L is considered indicative of functional B12 insufficiency (holoTC). This active fraction is a far more sensitive marker of cellular B12 status than total serum B12, which can appear normal even when tissues are starved.

The rs5749131 variant lies about 1.2–1.5 kb upstream of the TCN2 gene start site — squarely in the promoter and regulatory region that controls how much transcobalamin II your body produces. Carriers of the A allele show measurably reduced holoTC, suggesting the variant alters TCN2 transcriptional activity.

The Mechanism

Unlike the well-characterised TCN2 Pro259Arg missense variant (rs1801198), which changes the protein's B12-binding domain, rs5749131 sits in non-coding regulatory DNA. The precise mechanism has not yet been elucidated, but the upstream location suggests it may affect a transcription factor binding site or promoter element that governs TCN2 expression levels. Lower TCN2 expression would produce less transcobalamin protein, directly reducing the pool available to bind and transport B12 — the same endpoint reached by a different route from the missense variant.

The nearby variant rs5753231 (position 30,607,082, ~1.2 kb downstream of rs5749131) was identified as a secondary independent signal at the TCN2 locus in a large GWAS of serum B12, suggesting this regulatory region harbours at least two distinct functional elements controlling TCN2 expression.

The Evidence

A 2024 proteogenomic GWAS33 2024 proteogenomic GWAS
Western D et al. Proteogenomic analysis of human cerebrospinal fluid identifies neurologically relevant regulation and implicates causal proteins for Alzheimer's disease. Nat Genet, 2024 examining holotranscobalamin-2 levels in cerebrospinal fluid of 3,506 individuals identified rs5749131-A as strongly associated with reduced holoTC (p = 5.0×10⁻²⁴⁵, beta = −0.81 SD per A allele). While this was measured in CSF rather than serum, CSF holoTC reflects the same TCN2-mediated transport system and represents B12 delivery to neurological tissues specifically.

The TCN2 locus GWAS44 TCN2 locus GWAS
Grarup N et al. Genetic architecture of vitamin B12 and folate levels uncovered applying deeply sequenced large datasets. PLoS Genet, 2013 in 45,576 individuals identified multiple independent signals at the TCN2 locus associated with serum B12, with conditional analyses showing a secondary signal immediately 5′ to TCN2 — the same upstream region as rs5749131. This suggests variants in this regulatory region collectively influence TCN2 expression and B12 transport capacity.

A comprehensive pathway analysis55 comprehensive pathway analysis
Low HQ et al. A comprehensive association analysis of homocysteine metabolic pathway genes in Singaporean Chinese with ischemic stroke. PLoS One, 2011 studying 25 homocysteine pathway genes in 360 stroke patients and 360 controls identified rs5749131 among variants in TCN2 associated with ischemic stroke risk, consistent with the known link between impaired B12 transport, elevated homocysteine, and vascular disease.

Practical Implications

The practical implication is similar to that of the TCN2 missense variant (rs1801198): reduced holoTC may mean your cells receive less B12 than total serum measurements suggest. If you carry the A allele, requesting holotranscobalamin or methylmalonic acid (MMA) testing provides a truer picture of your cellular B12 status than standard total B12 panels.

Because this is a regulatory variant, strategies that maximise circulating B12 concentration — using bioavailable forms (methylcobalamin or hydroxocobalamin), sublingual delivery, and adequate dietary intake — can help maintain sufficient holoTC even if the variant modestly reduces transcobalamin expression.

The association with stroke in the Low et al. study is consistent with the well-established pathway: reduced holoTC → reduced cellular B12 → impaired homocysteine remethylation → elevated homocysteine → increased vascular risk. Monitoring homocysteine is a practical downstream check on whether B12 transport is functionally adequate.

Interactions

rs5749131 and rs1801198 both act on TCN2 but via different mechanisms — one regulatory, one structural. Carriers of risk alleles at both variants may have additive reductions in holoTC, though no published study has formally tested this combination.

Within the broader one-carbon cycle: methionine synthase (MTR, rs1805087) uses B12 as a cofactor to recycle homocysteine to methionine. Variants in MTR or MTRR (rs1801394) that reduce enzyme efficiency compound the effect of reduced B12 delivery. MTHFR C677T (rs1801133) variants impair the parallel folate arm of the cycle; combined impairment of both B12 delivery (TCN2) and folate processing (MTHFR) may produce the largest increases in homocysteine.

Nutrient Interactions

vitamin B12 altered_metabolism
holotranscobalamin reduced_absorption

Genotype Interpretations

What each possible genotype means for this variant.

GG Normal B12 Transport Regulation Normal

Common protective genotype — normal transcobalamin expression and holoTC levels

The GG genotype at rs5749131 represents the allele combination associated with higher holotranscobalamin in GWAS data. At the population level, G is the majority allele globally (58.4%) though this varies considerably by ancestry — G is predominant in African populations (~82%) but less common in East Asian populations (~25%).

Standard total serum B12 testing is a reasonable screen for individuals with this genotype, though holotranscobalamin remains a more precise functional marker for anyone.

AG Reduced Transport Regulation Intermediate

One copy of the A allele — mildly reduced holotranscobalamin

With one A allele and one G allele, your TCN2 expression regulation is intermediate. The GWAS association (beta = −0.81 per A allele in a CSF holoTC study at p = 5×10⁻²⁴⁵) suggests an additive dose effect: each A allele progressively reduces transcobalamin levels. One copy produces a modest reduction compared to GG, with the full effect seen in AA homozygotes.

Because total serum B12 captures both the metabolically active (holoTC) and inactive (haptocorrin-bound) fractions, your total B12 may appear normal while cellular delivery is modestly reduced.

AA Low Transport Regulation Reduced

Reduced transcobalamin expression — lower holotranscobalamin and potential functional B12 insufficiency

The AA genotype represents the homozygous form of the risk allele at rs5749131. The 2024 proteogenomic GWAS (PMID 39528825) found a beta of −0.81 SD per A allele for holotranscobalamin-2 levels (p = 5.0×10⁻²⁴⁵), meaning AA homozygotes have an approximately 1.6 SD lower holoTC than GG homozygotes — a substantial reduction.

Because this variant is regulatory (upstream of TCN2), not structural, it likely reduces the amount of transcobalamin II protein produced rather than impairing the protein's B12-binding function directly. The end result is the same: less transcobalamin available to load and transport B12, lower circulating holoTC, and reduced B12 delivery to tissues.

Total serum B12 can remain within the normal reference range while holoTC is depleted — the classic "hidden deficiency" pattern. Elevated methylmalonic acid (MMA) is the functional downstream marker confirming whether cellular B12 insufficiency is occurring.

The Low et al. stroke study in Singaporean Chinese (PMID 21935458) identified rs5749131 among TCN2 variants associated with ischemic stroke risk — consistent with the known pathway from reduced holoTC to impaired homocysteine remethylation to elevated homocysteine to increased vascular risk.