Age-related changes in expression of CTL2/SLC44A2 and its isoforms in the mouse inner ear

Abstract

The membrane glycoprotein CTL2/SLC44A2 is expressed by supporting cells in the inner ear and has been identified as a target of antibodies that may induce auto-immune hearing loss. To determine if CTL2/SLC44A2 also has roles in inner ear development and to distinguish between isoform-specific roles, we assessed age-related changes in expression of CTL2/SLC44A2 isoforms and protein in the developing murine inner ear. We determined that both isoform p1 and isoform p2 (named for the upstream p1 and proximal p2 promoters that control alternate exons 1a and 1b) were robustly expressed as early as E14 and persisted during embryonic development, but after birth the p1 isoform fell to barely detectable levels while isoform p2 levels were maintained. This trend continued and became even more apparent later in post-natal development and remained in mature ears until at least 6 weeks of age. In aged (18mo old) mice, the level of isoform p1 transcripts rose again to levels similar to the p2 isoform like that seen early in development. At the earliest stage examined, CTL2/SLC44A2 protein was expressed in both immature supporting cells and immature sensory cells, but after birth expression in the sensory cells declined in both the utricle and cochlea and by day P1 expression of CTL2/SLC44A2 was restricted to supporting cells. The changes we observed in isoform distribution are indicative of differential developmental roles and age related changes between the two isoforms of CTL2/SLC44A2 in the inner ear.

Hair Cell Loss, Spiral Ganglion Degeneration, and Progressive Sensorineural Hearing Loss in Mice with Targeted Deletion of Slc44a2/Ctl2

“…Immunohistochemistry was done using anti-myosin 7A (hair cells) and rhodamine phalloidin (actin) using published protocols (Sha et al 2008 ). Expression of Slc44a2 in the transgenic mice was carried out using affinity-purified rabbit anti-CTL2-NT as described previously (Beyer et al 2011 ). Inner ear cross sections were generated in the KHRI Core Histology Facility and evaluated.…”

Choline transporter‐like proteins CTLs/SLC44 family as a novel molecular target for cancer therapy

“…CTL2 mRNA and protein are expressed in human inner ear and rat renal epithelial cells . In addition, age‐related changes in the expression of CTL2 N ‐terminal isoforms have been reported in the murine inner ear . CTL2 is expressed as two isoforms (CTL2‐P1 and CTL2‐P2) in the cochlea, tongue, heart, colon, lung, kidney, liver and spleen, which suggests that tissue‐specific differences may influence its function in each tissue.…”

Full‐length nucleotide sequences of 30 common SLC44A2 alleles encoding human neutrophil antigen‐3

“…The SLC44A2 (solute carrier protein 44A2) gene encoding the CTL2 protein is located on chromosome 19p13.2 and consists of 22 exons. There are two isoforms of the full‐length cDNA, which differ only by the amino acids encoded by their respective first exon . The HNA‐3a/b antigens are expressed on the first extracellular loop of CTL2 protein and are caused by the nonsynonymous single‐nucleotide polymorphism (SNP) G>A in Exon 7 at Position 461 (rs2288904) in the mRNA Isoform 1 (NM_020428.3) leading to an amino acid substitution of Arg154 (HNA‐3a, SLC44A2*1 ) to Gln154 (HNA‐3b, SLC44A2*2 ) in Protein Isoform 1 (NP_065161.3) .…”

“…There are two isoforms of the full-length cDNA, which differ only by the amino acids encoded by their respective first exon. [11][12][13][14] The HNA-3a/b antigens are expressed on the first extracellular loop of CTL2 protein and are caused by the nonsynonymous single-nucleotide polymorphism (SNP) G>A in Exon 7 at Position 461 (rs2288904) in the mRNA Isoform 1 (NM_020428.3) leading to an amino acid substitution of Arg154 (HNA-3a, SLC44A2*1) to Gln154 (HNA-3b, SLC44A2*2) in Protein Isoform 1 (NP_065161.3). 10,15,16 The Leu153Phe (457C>T; rs147820753) has been reported to yield false-negative genotyping results for HNA-3a if the specific primer encompasses this variation.…”