SLC26A4/PDS gene 

The SLC26A4 gene consists of 21 exons located at the DFNB4 locus. It encodes the pendrin protein, which comprises 780 amino acids and is a multifunctional exchanger protein found in the thyroid, inner ear, and kidney. A lack of pendrin causes Pendred Syndrome. The SLC26A4 gene, in patients with Pendred Syndrome, the mutation of the gene leads to an intracellular ionic imbalance [10]. SLC26A4 is expressed by an Iodide/Chloride exchange in the thyroid [11] and Cl-/HCO3- exchange in the inner ear [12]. These exchanges occur during late embryonic and early postnatal development of the inner ear [13]. Pendrin also allows bicarbonate transport into the cochlear endolymph, which bathes the inner ear’s sensory cells to help convey information about sound, position, and balance [14]. Mutated pendrin may lead to iodide organification defects [15]. Most patients with PS have pathogenic variants and biallelic mutations in the SLC26A4 gene [8], [9]. Biallelic mutations of SLC26A4 are associated with abnormal iodide organification, increased thyroid gland volume, severe hearing loss, and bilateral enlarged vestibular aqueduct. Despite Pendred being characterized as an autosomal-recessive disorder, a single mutated allele of SLC26A4 presents with less severe consequences, and some may have normal iodide organification, normal thyroid gland volume, less severe hearing loss, and bilateral/unilateral EVA. The prevalence of a mutated SLC26A4 gene in PS patients is estimated at around 90%, which marks SLC26A4 as one of the most commonly mutated genes in  Pendred Syndrome [16].

SLC26A4 Effect on Thyroid

The thyroidal iodine organification defect common in PS results in the development of a goiter, which later becomes nodular [17]. Typically, potassium perchlorate transports iodide into thyroid folliculocytes across the basolateral membrane. However, in PS, high amounts of iodide are discharged in the thyroid. Despite Scott et al’s proposition that normal thyroid function in NSEVA (nonsyndromic enlarged vestibular aqueduct) patients is the consequence of residual activity encoded by mutated SLC26A4 variants [18], analysis of Ito et al [19] concluded that an enlarged thyroid and PS is dependent on the presence of 2 mutant SLC26A4 alleles, and NSEVA is associated with 1 or 0 mutant alleles [16], [20], [21].

SLC26A4 Effect on Auditory System

An Enlarged Vestibular Aqueduct (EVA) is a vestibular aqueduct exceeding 1.5 mm, measured between the common crus and external aperture. While Pendred Syndrome is characterized by the enlargement of the vestibular aqueduct, not all EVA patients have SLC26A4 mutations. In 50% of EVA populations, no mutations of SLC26A4 are detected [19]. Typically, Pendred Syndrome patients display cochlear hypoplasia secondary to a small or absent cochlear nerve, expansion of the scala media (cochlear duct), or an enlargement of the endolymphatic sac and duct [6], [22]–[24]. Due to additional oxidative stress, abnormal cell stretching, and impaired cell-to-cell communication in the stria vascularis, PS is associated with a reduced endocochlear potential [12], [25]–[27].

Hearing loss begins in the first few years of life, and is sensorineural or mixed, typically asymmetric, ranging from mild to profound. Sensorineural hearing loss occurs after inner ear damage. Mixed hearing loss occurs after damage in the outer or middle ear and in the inner ear or the nerve pathway to the brain. Hearing fluctuates downward following head trauma or barotrauma. In 92% of ears with SLC26A4 mutations, hearing loss fluctuation is observed, and some progressed at 1 decibel/year without environmental factors [16], [28], [29]. The type of mutation does not affect the severity of hearing loss, which is associated with the number of mutant alleles. Two mutant alleles result in more significant and fluctuating hearing loss when compared to those with one or zero mutant alleles [16], [20], [29], [30]. Hearing loss is also not associated with the severity of the enlarged vestibular aqueduct, suggesting that endolymphatic hydrops are not responsible for hearing loss [29], [31].

SLC26A4 Mutation Variants

8,647 different mutations of SLC26A4 have been reported, and 487 are classified as pathogenic [33]. The majority of mutations present in Pendred Syndrome are missense mutations, where mutant proteins are retained in the endoplasmic reticulum. Other possible mutations include nonsense mutations, splicing mutations, partial duplications, insertions, and deletions. Highly variable regions of mutations are exon 8, 19, 10, 17, and 15 [34]. These mutations affect iodide transport, which disrupts the protein function due to resulting iodide organification defects [15].  

Common mutations include E29Q, V138F, G209V, L236P, IVS8+1 G>A, R409H, T410M, T416P, Y78C, T193I, F355S, L445W, Y530H, S694P, D724N, 2127delT [16]. 

Variants in mutations also reflect ethnic differences. Three founder mutations in SLC26A4 have been identified in Caucasians, which are c.707T>C, c.1246A>C, and c.1001+ 1G>A mutations. The majority of mutations reported in China included the C.919-2 A>G mutation and C.2168A>G mutation. In South America and North America mutations C.1826T>G and C.1001 + 1G>A are more common, whereas C.2168A>G mutations are present in Koreans [35]. Recently, CEVA (Caucasian EVA) has been discovered to be a recessive mutant allele that is present in a pathogenic variant of SLC26A4, and is generally identified in Caucasian patients. CEVA includes 12 variants in introns or intergenic regions upstream of SLC26A4 [36].

In one case study, a patient was found with a compound heterozygosity variant in a mutated C.1174A>T. This affects amino acid position 535-729 in the STAS domain that affects protein function. In another patient with EVA, p.V690A, a missense mutation, was located on the same amino acid position, however there was no functional defect. One combination of compound heterozygosity (both parent genes harbor different mutations) present in SLC26A4 is the C.1341 + 1G>C mutation and the C.2069T>C, which are classified as disease-causing mutations, or DM. Using these classifications, enlarged vestibular aqueduct syndrome can be detected during neonatal hearing screening [16].

Ephrins and Pendrin

Reference: [33]

EphA2 is another gene implicated in the development of  PS. Typically, Pendrin is a binding partner of EPHA2. EPH receptors interact with plasma-membrane-bound ephrin ligands. Ephrins are categorized into two subclasses, A subclass (ligands for GPI-anchored EphA receptors) and B subclass (ligands for EphB1-6 tyrosine kinase receptors). EphB/ephrin-B2 is responsible for regulating vascular endothelial growth factor receptors such as VEGFR, which are responsible for transmembrane protein localization and compartmentalization of cells in epithelial tissue formation. Loss of ephrin-B2 results in abnormalities in the inner ear, disrupting cell proliferation, cell survival, folding of the endolymphatic epithelium, and abnormally formed otoconia [37]. 

As SLC26A4 is responsible for making Pendrin, a mutated SLC26A4 will impact protein EPHA2’s function. EphA2-null and ephrin-B2 deficient mice both exhibit abnormal structures in epithelial tissues and mislocalization of pendrin in the inner ear and thyroid [32]. However, stimulation of EphA2 and ephrin-B2 causes EphA2 and pendrin to move inside the cell from the outer membrane. This leads to a weaker self-activation of EphA2 compared to when activated with ephrin-A1. Due to EphA2’s inability to bind with ephrin-B2, it results in a failure of ephrin-B2 to induce internalization of pendrin. PS patients that bear a mono-allelic mutation of SLC26A4 will typically have point mutations that lead to amino acid substitution in EPHA2 [32].

EphA2 knock-out mice have an enlarged lumen, a decreased thickness of the stria vascularis, and a thyroid goiter, which are all present in PS patients [32]. EFNB2 (inner ear epithelial cell gene) encodes ephrin-B2 (responsible for the growth and morphogenesis of the endolymphatic sac and duct of the inner ear). EFNB2-deficient mice have vestibular-behavioral dysfunction and abnormal endolymph homeostasis, similar to Pendred symptoms [37].

Typically, EphA2 receptors are exclusively activated by ephrin-A; however, if  EphA2 is superimposed to EphA4 in a  complex with ephrin-B2, EphA2 gains the ability to bind to ephrin-B2 [37].

EPHA2 Mutations in Pendred Syndrome Patients

Missense mutations of the EphA2 gene responsible for Pendred syndrome patients include SLC26A4: c.1300G>A (p.434A>T), EPHA2: c.1063G>A (p.G355R) and SLC26A4: c.1229C>A (p.410T>M), EPHA2: c.1532C>T (p.T511M).