Mutations in OSBPL2 cause hearing loss associated with primary cilia defects via sonic hedgehog signaling
Hairong Shi, Hongshun Wang, Cheng Zhang, Yajie Lu, Jun Yao, Zhibin Chen, Guangqian Xing, Qinjun Wei, Xin Cao
Hairong Shi, Hongshun Wang, Cheng Zhang, Yajie Lu, Jun Yao, Zhibin Chen, Guangqian Xing, Qinjun Wei, Xin Cao
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Research Article Genetics Otology

Mutations in OSBPL2 cause hearing loss associated with primary cilia defects via sonic hedgehog signaling

Abstract

Defective primary cilia cause a range of diseases called ciliopathies, which include hearing loss (HL). Variants in the human oxysterol-binding protein like 2 (OSBPL2/ORP2) are responsible for autosomal dominant nonsyndromic HL (DFNA67). However, the pathogenesis of OSBPL2 deficiency has not been fully elucidated. In this study, we show that the Osbpl2-KO mice exhibited progressive HL and abnormal cochlear development with defective cilia. Further research revealed that OSBPL2 was located at the base of the kinocilia in hair cells (HCs) and primary cilia in supporting cells (SCs) and functioned in the maintenance of ciliogenesis by regulating the homeostasis of PI(4,5)P2 (phosphatidylinositol 4,5-bisphosphate) on the cilia membrane. OSBPL2 deficiency led to a significant increase of PI(4,5)P2 on the cilia membrane, which could be partially rescued by the overexpression of INPP5E. In addition, smoothened and GL13, the key molecules in the Sonic Hedgehog (Shh) signaling pathway, were detected to be downregulated in Osbpl2-KO HEI-OC1 cells. Our findings revealed that OSBPL2 deficiency resulted in ciliary defects and abnormal Shh signaling transduction in auditory cells, which helped to elucidate the underlying mechanism of OSBPL2 deficiency in HL.

Authors

Hairong Shi, Hongshun Wang, Cheng Zhang, Yajie Lu, Jun Yao, Zhibin Chen, Guangqian Xing, Qinjun Wei, Xin Cao

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Figure 6

Shh signal transduction was rescued by alleviating the accumulation of PI(4,5)P2 at the base of the cilia.

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Shh signal transduction was rescued by alleviating the accumulation of P...
(A) Immunofluorescence staining of Osbpl2–/– HEI-OC1 cells with anti-SMO (green), anti-acetylated tubulin (red), and DAPI (blue). In Osbpl2–/– HEI-OC1 cells expressing 5HT6-HA-INPP5E, the localization of ciliary SMO was partially rescued. Dashed frames denote the locally zoomed regions in solid frames (bottom right). Scale bar: 5 μm. (B) Quantification of ciliary SMO intensity in Osbpl2–/– HEI-OC1 cells with or without INPP5E expression (at least 30 cells from a microscope field, each dot represents a cell. **P < 0.01 by 2-tailed Student’s t test). (C) Immunofluorescence staining of Osbpl2–/– HEI-OC1 cells with anti-GLI3 (purple), anti-acetylated tubulin (green), and anti–gamma-tubulin (red). In Osbpl2–/– HEI-OC1 cells expressing 5HT6-HA-INPP5E, the localization of ciliary GLI3 was partially rescued. Dashed frames denote the locally zoomed regions in solid frames (bottom right). Scale bar: 5 μm. (D) Quantification of ciliary GLI3 intensity in Osbpl2–/– HEI-OC1 cells with or without INPP5E expression (at least 30 cells from a microscope field, each dot represents a cell; ***P < 0.001 by 2-tailed Student’s t test). (E) Schematic diagram of OSBPL2 regulating ciliogenesis and Shh signaling transduction in auditory cells. OSBPL2 was localized at the base of the cilia and regulated the homeostasis of ciliary PI(4,5)P2, which affected the ciliogenesis and thereby influenced transduction of the Shh signaling pathway. OSBPL2 deficiency led to dyshomeostasis of ciliary PI(4,5)P2, which was responsible for ciliary defects and inhibited Shh signal transduction.