A de novo silencer causes elimination of MITF-M expression and profound hearing loss in pigs

Lei Chen, Weiwei Guo, Lili Ren, Mingyao Yang, Yaofeng Zhao, Zongyi Guo, Haijin Yi, Mingzhou Li, Yiqing Hu, Xi Long, Boyuan Sun, Jinxiu Li, Suoqiang Zhai, Tinghuan Zhang, Shilin Tian, Qingyong Meng, Ning Yu, Dan Zhu, Guoqing Tang, Qianzi Tang, Liming Ren, Ke Liu, Shihua Zhang, Tiandong Che, Zhengquan Yu, Nan Wu, Lan Jing, Ran Zhang, Tao Cong, Siqing Chen, Yiqiang Zhao, Yue Zhang, Xiaoqing Bai, Ying Guo, Lidong Zhao, Fengming Zhang, Hui Zhao, Liang Zhang, Zhaohui Hou, Jiugang Zhao, Jianan Li, Lijuan Zhang, Wei Sun, Xiangang Zou, Tao Wang, Liangpeng Ge, Zuohua Liu, Xiaoxiang Hu, Jingyong Wang, Shiming Yang, Ning Li

Genesis of novel gene regulatory modules is largely responsible for morphological and functional evolution. De novo generation of novel cis-regulatory elements (CREs) is much rarer than genomic events that alter existing CREs such as transposition, promoter switching or co-option. Only one case of de novo generation has been reported to date, in fish and without involvement of phenotype alteration. Yet, this event likely occurs in other animals and helps drive genetic/phenotypic variation. Using a porcine model of spontaneous hearing loss not previously characterized we performed gene mapping and mutation screening to determine the genetic foundation of the phenotype. We identified a mutation in the non-regulatory region of the melanocyte-specific promoter of microphthalmia-associated transcription factor (MITF) gene that generated a novel silencer. The consequent elimination of expression of the MITF-M isoform led to early degeneration of the intermediate cells of the cochlear stria vascularis and profound hearing loss, as well as depigmentation, all of which resemble the typical phenotype of Waardenburg syndrome in humans. The mutation exclusively affected MITF-M and no other isoforms. The essential function of Mitf-m in hearing development was further validated using a knock-out mouse model. Elimination of the MITF-M isoform alone is sufficient to cause deafness and depigmentation. To our knowledge, this study provides the first evidence of a de novo CRE in mammals that produces a systemic functional effect.