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Examination of phosducin gene

磷酸蛋白基因检查

基本信息

批准号：
08672004
负责人：
ABE Toshiaki
金额：
$ 1.6万
依托单位：
Tohoku University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
1996
资助国家：
日本
起止时间：
1996 至 1997
项目状态：
已结题

项目摘要

Phosducin inhibit phototransduction signal by interacting with transducin betagamma-subunit (1) by cAMP-dependent protein kinase A,but phosducin does not inhibit the initial onset of phototransduction signal (2) (3). The gene was suspected to have gene family from the Southern blotting analysis (4) and was located close to the disease locus of usher syndrome type II (5), (6). We screened the bovine cDNA library using the probe, which was generated the rat retina cDNA by polymerase chain reaction. We cloned a gene, which was considered bovine phosducin like protein as that of rat phosducin like protein reported by Miles and coworkers (7). The gene produced isoforms by alternative splicing and interestingly, one of the form was extensively expressed in the rat retina.We constracted transgenic mice harvaring mutant phosducin gene, which had isoleucin instead of serin at amino acid 73. The transgenic mice showed 50% recovery of the b-wave of electroretinogram after light stimulation as that of normal control mice under light adaptation. The transgenic mice also showed 50% recovery under dark adaptation. We speculated that phosducin may play an important role for the photoreceptor adaptation.Microscopic and electronmicroscopic examination revealed that photoreceptor cells show normal appearance at the early post natal days, but gradually decreased the length of the outer segment and outer nuclear layr.Immunohistochemical study revealed that immunostaining was also observed at habenula commissura amygdala or superiorcolliculus.

Phosducin通过cAMP依赖性蛋白激酶A与转导素β γ亚基（1）相互作用抑制光转导信号，但Phosducin不抑制光转导信号的初始启动（2）（3）。从Southern印迹分析中怀疑该基因具有基因家族（4），并且位于接近于II型usher综合征的疾病位点（5），（6）。以大鼠视网膜cDNA为探针，通过聚合酶链反应筛选牛视网膜cDNA文库。我们克隆了一个基因，该基因被认为是牛类胰蛋白酶样蛋白，与Miles等报道的大鼠类胰蛋白酶样蛋白相同（7）。该基因通过选择性剪接产生异构体，有趣的是，其中一种形式在大鼠视网膜中广泛表达。我们复制了获得突变体Adducin基因的转基因小鼠，该基因在氨基酸73处具有异亮氨酸而不是丝氨酸。转基因小鼠在光适应条件下，光刺激后视网膜电图b波恢复正常对照小鼠的50%。转基因小鼠在暗适应下也显示出50%的恢复。光镜和电镜观察表明，光感受器细胞在纳塔尔后早期外观正常，但外节和外核层的长度逐渐变短，免疫组化研究表明，在缰连合、杏仁核和上丘也有免疫染色。