Bost-Usinger L, Chen R J, Hillman D, Park H, Burnside B
Department of Molecular and Cell Biology, University of California, Berkeley 94720-3200, USA.
Exp Eye Res. 1997 May;64(5):781-94. doi: 10.1006/exer.1996.0271.
Kinesins comprise a large superfamily of microtubule-based motor proteins, individual members of which mediate specific types of motile processes. To identify kinesin family members (KIFs) that are critical to retinal function and thus to vision, a reverse transcriptase polymerase chain reaction (RT-PCR) cloning strategy was used to isolate putative KIFs expressed in the neural retina and retinal pigmented epithelium (RPE) of the striped bass, Morone saxatilus. Eleven fish KIFs (FKIFs) were isolated from neural retina and six of the same FKIFs were also isolated from RPE. One of the KIFs identified in this screen, FKIF2, was the most prevalent clone detected both in the retina (41% of clones) and RPE (72% of clones). Based on predicted amino acid sequence homology within the motor domain, seven of the FKIFs have been tentatively assigned to known kinesin families: the kinesin heavy chain family (FKIF1, 5 and 9), the unc104/KIF1 family (FKIF3 and 8), the KIF2 family (FKIF4), and the cKIF family (FKIF2). Northern blot analysis revealed that each detectable FKIF exhibited a unique tissue-specific expression pattern. Since FKIF2 was more highly expressed in retina than in any other tissue tested, including brain, and was the most abundant KIF message expressed in both retina and RPE, it was examined in more detail and the complete approximately 2.3 kb open reading frame for FKIF2 was cloned and sequenced. The predicted amino acid sequence indicates that FKIF2 has a C-terminal motor domain, and thus is a member of the cKIF family. FKIF2 is only 36.5% identical at the amino acid level to the most closely related cKIF in the database, suggesting that FKIF2 may be a novel member of this family. Antibodies raised against a unique peptide specific to FKIF2 recognize an approximately 80 kd protein in homogenates of retina, RPE, brain and kidney. The pronounced expression of FKIF2 in retina and RPE suggests that FKIF2 may play an important role in microtubule-dependent motile events in these two tissues.
驱动蛋白构成了一个基于微管的运动蛋白的大型超家族,其各个成员介导特定类型的运动过程。为了鉴定对视网膜功能乃至视觉至关重要的驱动蛋白家族成员(KIFs),采用逆转录聚合酶链反应(RT-PCR)克隆策略,从条纹鲈(Morone saxatilus)的神经视网膜和视网膜色素上皮(RPE)中分离出假定的KIFs。从神经视网膜中分离出11种鱼类KIFs(FKIFs),同时从RPE中也分离出6种相同的FKIFs。在此筛选中鉴定出的一种KIF,即FKIF2,是在视网膜(占克隆的41%)和RPE(占克隆的72%)中检测到的最普遍的克隆。基于运动结构域内预测的氨基酸序列同源性,7种FKIFs已被初步归入已知的驱动蛋白家族:驱动蛋白重链家族(FKIF1、5和9)、unc104/KIF1家族(FKIF3和8)、KIF2家族(FKIF4)和cKIF家族(FKIF2)。Northern印迹分析表明,每个可检测到的FKIF都表现出独特的组织特异性表达模式。由于FKIF2在视网膜中的表达高于包括脑在内的任何其他测试组织,并且是在视网膜和RPE中表达最丰富的KIF信息,因此对其进行了更详细的研究,并克隆和测序了FKIF2完整的约2.3 kb开放阅读框。预测的氨基酸序列表明FKIF2具有C末端运动结构域,因此是cKIF家族的成员。FKIF2在氨基酸水平上与数据库中关系最密切的cKIF仅36.5%相同,这表明FKIF2可能是该家族的一个新成员。针对FKIF2特有的独特肽段产生的抗体,在视网膜、RPE、脑和肾的匀浆中识别出一种约80 kd的蛋白质。FKIF2在视网膜和RPE中的显著表达表明,FKIF2可能在这两个组织中依赖微管的运动事件中发挥重要作用。