The role of CDHR3 in susceptibility to otitis media

Abstract

Otitis media (OM) is common in young children and can cause hearing loss and speech, language, and developmental delays. OM has high heritability; however, little is known about OM-related molecular and genetic processes. CDHR3 was previously identified as a locus for OM susceptibility, but to date, studies have focused on how the CDHR3 p.Cys529Tyr variant increases epithelial binding of rhinovirus-C and risk for lung or sinus pathology. In order to further delineate a role for CDHR3 in OM, we performed the following: exome sequencing using DNA samples from OM-affected individuals from 257 multi-ethnic families; Sanger sequencing, logistic regression and transmission disequilibrium tests for 407 US trios or probands with OM; 16S rRNA sequencing and analysis for middle ear and nasopharyngeal samples; and single-cell RNA sequencing and differential expression analyses for mouse middle ear. From exome sequence data, we identified a novel pathogenic CDHR3 splice variant that co-segregates with OM in US and Finnish families. Additionally, a frameshift and six missense rare or low-frequency variants were identified in Finnish probands. In US probands, the CDHR3 p.Cys529Tyr variant was associated with the absence of middle ear fluid at surgery and also with increased relative abundance of Lysobacter in the nasopharynx and Streptomyces in the middle ear. Consistent with published data on airway epithelial cells and our RNA-sequence data from human middle ear tissues, Cdhr3 expression is restricted to ciliated epithelial cells of the middle ear and is downregulated after acute OM. Overall, these findings suggest a critical role for CDHR3 in OM susceptibility. KEY MESSAGES: • Novel rare or low-frequency CDHR3 variants putatively confer risk for otitis media. • Pathogenic variant CDHR3 c.1653 + 3G > A was found in nine families with otitis media. • CDHR3 p.Cys529Tyr was associated with lack of effusion and bacterial otopathogens. • Cdhr3 expression was limited to ciliated epithelial cells in mouse middle ear. • Cdhr3 was downregulated 3 h after infection of mouse middle ear.

Keywords: CDHR3; Expression; Microbiome; Middle ear; Otitis media; rs6967330.

Figure 1

Three European-descent families with co-segregation of the same CDHR3 splice variant c.1653+3G>A (rs117797654) variant with otitis media (OM). Black symbols, OM-affected; clear symbols, no diagnosis of OM. Note that parents were ascertained for OM as adults, which does not completely rule out OM in early childhood. Alternatively, the variant may be inherited in an autosomal dominant pattern with reduced penetrance. Five additional Finnish probands (from two trios and three parent-proband pairs) are heterozygous for this splice variant (Table 1). A Minnesotan family UMN48 has three out of four OM-affected relatives that are heterozygous for the CDHR3 splice variant, however family UMN48 also has the known PLG variant c.112A>G (p.Lys38Glu, rs73015965) co-segregating with otitis media [32]. Probands from eight of the nine families with the splice variant, including the four families in the figure, have both RAOM and COME diagnoses

Figure 2

Relative taxa abundance in the middle ear (ME) and nasopharynx (NP). Left panels summarize average relative abundances (RA) of predominant bacterial genera (taxa with RA <0.5% were collapsed into the “Other” group). Right panels show the results of non-parametric Wilcoxon rank sum tests, which plot -log(nominal-p-values) for each genus along the x-axes. Blue lines indicate taxa that were of greater RA in the microbial samples from wildtype individuals vs. carriers of the CDHR3 p.Cys529Tyr variant, whereas red lines indicate taxa that were of greater RA in carriers of the p.Cys529Tyr variant compared with wildtype. Horizontal lines mark p-values equal to 0.1, 0.05, or 0.01. The CDHR3 p.Cys529Tyr variant was nominally associated with increased relative abundance of Streptomyces (p=0.04) in the ME and Lysobacter (p=0.01) in the NP

Figure 3

Relative Cdhr3 expression after inoculation of wildtype mouse middle ears with non-typeable Haemophilus influenzae (NTHi). Cdhr3 is significantly downregulated three hours post-infection (Table 5)