Coordinated control of connexin 26 and connexin 30 at the regulatory and functional level in the inner ear
Coordinated control of connexin 26 and connexin 30 at the regulatory and functional level in the inner ear
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DOI：
10.1073/pnas.0800831105
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发表时间：
2008-12-02
影响因子：
11.1
通讯作者：
Chiorini, John A.
中科院分区：
文献类型：
--
作者：
Ortolano, Saida;Di Pasquale, Giovanni;Chiorini, John A.
连接蛋白26 （Cx26）和连接蛋白30 （COO）由两个基因（分别为GJB2和GJa6）编码，这两个基因位于相同的复杂耳聋位点DFNB1的50 kb范围内。免疫细胞化学和定量PCR分析显示，Cx26在位于外毛细胞和血管纹之间的非感觉细胞中的蛋白水平和mRNA水平均下调。为了探索连接蛋白的协同调节，我们使用牛腺相关病毒操纵基因表达。通过牛腺相关病毒转导Cx30在Cx30 KO小鼠中过表达，可以恢复Cx26的表达，允许形成功能性的间隙连接通道，并恢复传播的Ca2+信号。用Cre重组酶载体转导Cx26（loxP/loxP）培养物，导致Cx26同时下调，细胞间通讯受损。Cx26和Cx30表达的协调调节似乎是通过PLC和NF-kappa B途径信号传导的结果，因为用20 nM ATP刺激P2Y受体20分钟，激活ip3介导的Ca2+反应，增加了Cx30 KO培养物中Cx26转录物的水平。这种效应通过表达一种稳定形式的I κ B抑制蛋白来抑制，该蛋白可以阻止nf - κ B的激活/易位。因此，我们的数据揭示了与遗传性耳聋相关的内耳连接蛋白表达的Ca2+依赖性控制，以及迄今为止无法解释的观察结果，即一些耳聋相关的DFNB1等位基因的特征是GJB2和GJB6表达的遗传减少。
Connexin 26 (Cx26) and connexin 30 (COO) are encoded by two genes (GJB2 and GJa6, respectively) that are found within 50 kb in the same complex deafness locus, DFNB1. Immunocytochemistry and quantitative PCR analysis of Cx30 KO mouse cultures revealed that Cx26 is downregulated at the protein level and at the mRNA level in nonsensory cells located between outer hair cells and the stria vascularis. To explore connexin coregulation, we manipulated gene expression using the bovine adeno-associated virus. Overexpression of Cx30 in the Cx30 KO mouse by transduction with bovine adeno-associated virus restored Cx26 expression, permitted the formation of functional gap junction channels, and rescued propagating Ca2+ signals. Ablation of Cx26 by transduction of Cx26(loxP/loxP) cultures with a Cre recombinase vector caused concurrent downregulation of Cx30 and impaired intercellular communication. The coordinated regulation of Cx26 and Cx30 expression appears to occur as a result of signaling through PLC and the NF-kappa B pathway, because activation of IP3-mediated Ca2+ responses by stimulation of P2Y receptors for 20 min with 20 nM ATP increased the levels of Cx26 transcripts in Cx30 KO cultures. This effect was inhibited by expressing a stable form of the I kappa B repressor protein that prevents activation/translocation of NF-kappa B. Thus, our data reveal a Ca2+-dependent control in the expression of inner ear connexins implicated in hereditary deafness as well as insight into the hitherto unexplained observation that some deafness-associated DFNB1 alleles are characterized by hereditable reduction of both GJB2 and GJB6 expression.