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Alternative splicing plays a pivotal role in modulating the function of eukaryotic proteins. In the inner ear, many genes undergo alternative splicing, and errors in this process lead to hearing loss. Cadherin 23 (CDH23) forms part of the so-called tip links, which are indispensable for mechanoelectrical transduction (MET) in the hair cells. Cdh23 gene contains 69 exons, and exon 68 is subjected to alternative splicing. Exon 68 of the Cdh23 gene is spliced into its mRNA only in a few cell types including hair cells. The mechanism responsible for the alternative splicing of Cdh23 exon 68 remains elusive. In the present work, we performed a cell-based screening to look for splicing factors that regulate the splicing of Cdh23 exon 68. RBM24 and RBM38 were identified to enhance the inclusion of Cdh23 exon 68. The splicing of Cdh23 exon 68 is affected in Rbm24 knockdown or knockout cells. Moreover, we also found that PTBP1 inhibits the inclusion of Cdh23 exon 68. Taken together, we show here that alternative splicing of Cdh23 exon 68 is regulated by RBM24, RBM38, and PTBP1. [ABSTRACT FROM AUTHOR]

PDZD7 is an important deafness gene, whose mutations are associated with syndromic and nonsyndromic hearing loss. PDZD7 contains multiple PDZ domains that are essential for organizing various proteins into protein complex. Several PDZD7-binding proteins have been identified, including usherin, ADGRV1, whirlin, harmonin, SANS and MYO7A, all belonging to USH proteins. Here, we report the identification of novel PDZD7-binding partners through yeast two-hybrid screening using the first two PDZ domains of PDZD7 as bait. Eleven proteins were identified, most of which have not been reported as PDZD7-binding partners before. Among the identified proteins, ADGRV1, gelsolin, and ß-catenin have been shown to play important roles in hearing, whereas the functions of other proteins in the inner ear remain elusive. We confirmed the expression of one candidate PDZD7-binding protein, CADM1, in the mouse inner ear and evaluated the auditory function of Cadm1 knockout mice by performing auditory brainstem response (ABR) measurement. Unexpectedly, Cadm1 knockout mice show normal hearing threshold, which might be explained by the possible compensation by its homologs that are also expressed in the inner ear. Taken together, our work identified several novel PDZD7-binding proteins, which will help us to further understand the role of PDZD7 in hearing transduction. [ABSTRACT FROM AUTHOR]

Protocadherin 15 (PCDH15) is a core component of hair cell tip-links and crucial for proper function of inner ear hair cells. Mutations of PCDH15 gene cause syndromic and nonsyndromic hearing loss. At present, the regulatory mechanisms responsible for the intracellular transportation of PCDH15 largely remain unknown. Here we show that PIST, a Golgi-associated, PDZ domain-containing protein, interacts with PCDH15. The interaction is mediated by the PDZ domain of PIST and the C-terminal PDZ domain-binding interface (PBI) of PCDH15. Through this interaction, PIST retains PCDH15 in the trans-Golgi network (TGN) and reduces the membrane expression of PCDH15. We have previously showed that PIST regulates the membrane expression of another tip-link component, cadherin 23 (CDH23). Taken together, our finding suggests that PIST regulates the intracellular trafficking and membrane targeting of the tip-link proteins CDH23 and PCDH15. [ABSTRACT FROM AUTHOR]