Severe piebaldism and profound hearing loss in mutant pigs derived from ENU mutagenesis
During N-ethyl-N-nitrosourea (ENU) mutagenesis, we designed a three-generation breeding scheme to retrieve dominant and recessive mutations in the F1 offspring(Hai, Cao, et al., 2017). A dominant mutation was identified in a male pig (G1-016105) with pigmentation abnormalities. The G1-016105 was mated with a wild-type Bama sow, resulting in 63 offspring from 10 litters (Fig. 1a). Among them, 27 offspring had the “two-ends” black color, and 36 showed severe piebaldism similar to G1-016105, including 16 males and 20 females (Fig. 1b). The mutant piglets showed symmetrical pigmentation abnormalities with white hair and skin on the forehead, cheek, the entire ear, and the distal part of the tail, which is highly similar to the distribution of piebaldism in humans (Fig. 1c). Moreover, the iris color of mutant pigs was normal. The blood cell analysis of peripheral blood did not find any marked difference in the mutant pigs (Table. S1).
As the hypopigmentation in the skin is primarily associated with hearing loss in human Waardenburg syndrome (WS)(Saleem, 2019), we conducted an auditory brainstem response (ABR) test in the mutant and wild-type littermates at postnatal day 7. Mutant pigs did not produce any recognizable waveforms up to 100 dB sound pressure level (SPL) stimuli in the range of 1–32 kHz, whereas wild-type pigs produced ABR thresholds at 10–30 dB SPL (Fig. 1d, 1e). The ABR test on every offspring showed a linkage between piebaldism and hearing loss. The frequency of the two phenotypes was compared with expected Mendelian ratios using a χ2 test (χc2=1.0159<χ0.05(1)2), suggesting that the mutant phenotype is inherited in an autosomal dominant pattern (Table 1).