Hildebrand Michael S, de Silva Michelle G, Klockars Tuomas, Rose Elizabeth, Price Margaret, Smith Richard J H, McGuirt Wyman T, Christopoulos Helen, Petit Christine, Dahl Hans-Henrik M
Department of Gene Identification and Expression, Murdoch Childrens Research Institute, Royal Children's Hospital, Flemington Road, Parkville, Vic. 3052, Australia.
Hear Res. 2004 Apr;190(1-2):149-60. doi: 10.1016/S0378-5955(04)00015-2.
Within the cochlea, the hair cells detect sound waves and transduce them into receptor potential. The molecular architecture of the highly specialised cochlea is complex and until recently little was known about the molecular interactions which underlie its function. It is now clear that the coordinated expression and interplay of hundreds of genes and the integrity of cochlear cells regulate this function. It was hypothesised that transcripts expressed highly or specifically in the cochlea are likely to have important roles in normal hearing. Microarray analyses of the Soares NMIE library, consisting of 1536 cDNA clones isolated from the mouse inner ear, suggested that the expression of the mechanoreceptor DRASIC was enriched in the cochlea compared to other tissues. This amiloride-sensitive ion channel is a member of the DEG/ENaC superfamily and a potential candidate for the unidentified mechanoelectrical transduction channel of the sensory hair cells of the cochlea. The cochlear-enriched expression of amiloride-sensitive cation channel 3 (ACCN3) was confirmed by quantitative real-time polymerase chain reaction. Using in situ hybridisation and immunofluorescence, DRASIC expression was localised to the cells and neural fibre region of the spiral ganglion. DRASIC protein was also detected in cells of the organ of Corti. DRASIC may be present in cochlear hair cells as the ACCN3 transcript was shown to be expressed in immortalised cell lines that exhibit characteristics of hair cells. The normal mouse ACCN3 cDNA and an alternatively spliced transcript were elucidated by reverse transcription polymerase chain reaction from mouse inner ear RNA. This transcript may represent a new protein isoform with an as yet unknown function. A DRASIC knockout mouse model was tested for a hearing loss phenotype and was found to have normal hearing at 2 months of age but appeared to develop hearing loss early in life. The human homologue of ACCN3, acid-sensing ion channel 3, maps to the same chromosomal region as the autosomal recessive hearing loss locus DFNB13. However, we did not detect mutations in this gene in a family with DFNB13 hearing loss.
在耳蜗内，毛细胞检测声波并将其转化为感受器电位。高度特化的耳蜗的分子结构很复杂，直到最近，对其功能背后的分子相互作用仍知之甚少。现在很清楚，数百个基因的协调表达和相互作用以及耳蜗细胞的完整性调节着这种功能。据推测，在耳蜗中高度表达或特异性表达的转录本可能在正常听力中发挥重要作用。对由从小鼠内耳分离的1536个cDNA克隆组成的索尔斯NMIE文库进行微阵列分析表明，与其他组织相比，机械感受器DRASIC在耳蜗中的表达更为丰富。这种对阿米洛利敏感的离子通道是DEG/ENaC超家族的成员，是耳蜗感觉毛细胞未确定的机械电转导通道的潜在候选者。通过定量实时聚合酶链反应证实了对阿米洛利敏感的阳离子通道3（ACCN3）在耳蜗中的高表达。使用原位杂交和免疫荧光技术，将DRASIC的表达定位到螺旋神经节的细胞和神经纤维区域。在柯蒂氏器的细胞中也检测到了DRASIC蛋白。由于ACCN3转录本在表现出毛细胞特征的永生化细胞系中表达，因此DRASIC可能存在于耳蜗毛细胞中。通过逆转录聚合酶链反应从小鼠内耳RNA中阐明了正常小鼠ACCN3 cDNA和一个选择性剪接的转录本。这个转录本可能代表一种功能未知的新蛋白质异构体。对一个DRASIC基因敲除小鼠模型进行听力损失表型测试，发现其在2个月大时听力正常，但似乎在生命早期就出现了听力损失。ACCN3的人类同源物，即酸敏感离子通道3，定位于与常染色体隐性听力损失位点DFNB13相同的染色体区域。然而，在一个患有DFNB13听力损失的家族中，我们没有检测到该基因的突变。
