rs137853000 — TMPRSS3 p.Arg216Leu (R216L)

TMPRSS3 p.Arg216Leu — A Pathogenic Missense Variant That Abolishes Serine Protease Activation

The TMPRSS3 gene encodes a type II transmembrane serine protease¹¹ type II transmembrane serine protease
Anchored to the cell membrane with its catalytic domain facing the extracellular space; expressed in cochlear inner hair cells, outer hair cells, spiral ganglion neurons, and the stria vascularis essential for the survival and maturation of cochlear hair cells. TMPRSS3 is unusual among serine proteases in that it undergoes autocatalytic activation — the full-length single-chain zymogen must cleave itself at a specific arginine residue to release the active catalytic domain. The p.Arg216Leu substitution strikes this cleavage site directly, converting the critical arginine to leucine and locking the protein permanently in its inactive zymogen form.

This variant was identified in two Turkish brothers with non-syndromic deafness born to consanguineous parents²² identified in two Turkish brothers with non-syndromic deafness born to consanguineous parents
Wattenhofer M et al., Human Genetics 2005, screening 25 Turkish families; R216L found homozygous in the affected siblings; the mutation was the highest-scoring TMPRSS3 mutation for LOD score linkage to chr21q22.3 in that cohort. It is catalogued in ClinVar (variation 4946) as pathogenic for autosomal recessive nonsyndromic hearing loss 8 (DFNB8), with OMIM allelic entry 605511.0005.

The Mechanism

TMPRSS3 is synthesized as an inactive single-chain precursor (zymogen). Activation requires autocatalytic cleavage at arginine 216³³ arginine 216
The canonical serine protease activation motif contains a conserved arginine or lysine immediately preceding the catalytic domain; cleavage here releases the prodomain and exposes the active-site triad, which generates the active two-chain form held together by a disulfide bond. This processing step is obligatory for all downstream activity.

The p.Arg216Leu substitution replaces the cleavage-site arginine with leucine — an amino acid that the protease cannot recognize as a cleavage substrate. Functional characterization in Xenopus oocytes⁴⁴ Functional characterization in Xenopus oocytes
Wattenhofer 2005 used the well-validated oocyte expression system to measure both autocatalytic processing and ENaC activation; neither occurred with R216L TMPRSS3 confirmed that the R216L mutant protein: (1) fails to undergo proteolytic self-cleavage, and (2) is completely unable to activate the epithelial sodium channel (ENaC). This distinguishes R216L from hypomorphic missense variants (such as p.Ala306Thr, which retains partial activity) — R216L produces a functional null.

The downstream cochlear consequence is hair cell degeneration. In mouse models, Tmprss3-deficient cochleae show normal morphology at birth but undergo rapid progressive hair cell loss beginning at postnatal day 12⁵⁵ Tmprss3-deficient cochleae show normal morphology at birth but undergo rapid progressive hair cell loss beginning at postnatal day 12
The onset coincides exactly with the moment hearing activates in mice; loss begins in the high-frequency basal cochlear turn and sweeps toward the apex, consistent with the down-sloping audiogram seen in human DFNB8 — the same postnatal timing when the organ of Corti first becomes mechanically and electrically active.

The Evidence

Functional evidence is conclusive. The 2005 Wattenhofer study provided direct biochemical proof that R216L abolishes TMPRSS3 activity through loss of autocatalytic processing. This is one of only a handful of TMPRSS3 variants with complete functional nullification confirmed in a heterologous expression system⁶⁶ This is one of only a handful of TMPRSS3 variants with complete functional nullification confirmed in a heterologous expression system
Most TMPRSS3 missense variants have not been functionally characterized; R216L and p.Ala306Thr are the best-characterized variants in the literature.

Population context. TMPRSS3 accounts for approximately 11% of autosomal recessive nonsyndromic hearing loss in Turkish populations⁷⁷ TMPRSS3 accounts for approximately 11% of autosomal recessive nonsyndromic hearing loss in Turkish populations
Compared to <1% in Caucasians, 0.7% in Japanese, 3% in Pakistani, 4.6% in Chinese, 5–6% in Tunisian, and 5.9% in Korean populations — Turkish prevalence is the highest documented, making it the highest-prevalence population for this gene. The R216L allele was specifically characterized in a Turkish family, and given the high TMPRSS3 burden in Turkish populations, this allele likely contributes to a meaningful fraction of Turkish ARNSHL cases. In gnomAD, the A allele at chr21:42383168 (the R216L plus-strand allele) appears at a global frequency of approximately 0.003% — consistent with a rare recessive pathogenic allele maintained at low frequency by heterozygous carrier state.

Genotype-phenotype correlation. A comprehensive genotype-phenotype analysis⁸⁸ comprehensive genotype-phenotype analysis
Nisenbaum et al. Audiology & Neurotology 2023, reviewing all published TMPRSS3 cases with natural history data established that biallelic severe/null TMPRSS3 alleles produce prelingual profound deafness (DFNB10), while combinations of one null allele with one missense allele typically produce postlingual progressive hearing loss (DFNB8) with a characteristic down-sloping audiogram progressing at approximately 0.3 dB per year⁹⁹ 0.3 dB per year
This rate applies to DFNB8 phenotype; DFNB10 patients present with congenital profound deafness and show essentially no residual cochlear function by audiometric testing. Because R216L is a functional null, homozygous R216L individuals would be expected to manifest DFNB10 (congenital profound deafness), while compound heterozygotes carrying R216L on one allele and a milder missense variant on the other might present with DFNB8 progressive loss.

Cochlear implantation outcomes in TMPRSS3-related hearing loss are excellent and consistent across populations¹⁰¹⁰ excellent and consistent across populations
International cohort of 127 patients, 16 centers, 6 countries; mean word recognition score 76% at follow-up. Age at implantation — not genotype — is the principal predictor of speech recognition outcome, with each year of delay associated with a measurable decrement in outcome. This makes early diagnosis and timely implantation the most critical clinical interventions.

Practical Implications

For confirmed homozygous (AA) individuals, this information directly supports early hearing intervention. For heterozygous (AC) carriers, hearing is expected to be normal but carrier status is clinically important for reproductive planning, particularly given the elevated TMPRSS3 carrier frequency in Turkish and other Middle Eastern populations.

For any individual with this variant, standard hearing health practices do not require modification — TMPRSS3-related hearing loss does not respond to dietary, supplement, or pharmacological interventions. The clinical value is entirely in diagnosis (confirming the cause of hearing loss in affected individuals), prognosis (predicting progression), and reproductive counseling (assessing offspring risk in carrier couples).

Interactions

The primary interaction is with other TMPRSS3 pathogenic alleles. Biallelic TMPRSS3 mutations can arise from homozygosity (two copies of the same allele, as in the original Turkish family carrying two R216L copies) or compound heterozygosity (one copy of R216L on one chromosome and a different pathogenic allele on the other). Given the diversity of TMPRSS3 pathogenic alleles documented across populations — including rs727503493 (c.208delC frameshift) and rs10421919 locus variants — comprehensive TMPRSS3 panel sequencing is required to identify compound heterozygotes, who will have two different pathogenic alleles.

A possible digenic interaction with GJB2¹¹¹¹ A possible digenic interaction with GJB2
GJB2 encodes connexin 26, the most common cause of ARNSHL in Caucasians (c.35delG allele, rs80338939); initial case reports suggested TMPRSS3/GJB2 digenic inheritance, later disputed by larger series has been proposed but is not confirmed. In clinical practice, when a single TMPRSS3 pathogenic allele is found in an individual with hearing loss, comprehensive deafness gene panel testing — including GJB2 — is warranted.