Zhang Houbin, Huang Wei, Zhang Haikun, Zhu Xuemei, Craft Cheryl M, Baehr Wolfgang, Chen Ching-Kang
Department of Neurobiology and Anatomy, University of Utah, Salt Lake City, USA.
Mol Vis. 2003 Jun 9;9:231-7.
The light-dependent redistribution of phototransduction components in photoreceptor cells plays a role in light adaptation. Upon illumination, rod and cone arrestins (Arr and cArr) translocate from the inner to the outer segments while transducin subunits (Talpha, Tbetagamma) translocate in the opposite direction. The underlying translocation mechanisms are unclear. This study examines these previously demonstrated translocation in mice with defective phototransduction.
The distribution of Arr, cArr, Talpha, and Tbetagamma was examined using immunoblotting and immunocytochemistry in dark- and light-adapted single knockout mice lacking G-protein coupled receptor kinase 1 (Grk1-/-) and double knockout mice lacking GRK1 and transducin alpha subunit (Grk1-/-/Gnat1-/-), or lacking GRK1 and arrestin (Grk1-/-/Arr-/-).
Arr redistributed in the light to the outer segments in Grk1-/- mice as well as in Grk1-/-/Gnat1-/- double knockout retinas. Immunoblotting revealed that approximately 25-50% of Arr associated with the membrane in light-adapted wild-type, Grk1-/- and Gnat1-/-/Grk1-/- mouse retinas. In contrast, cArr did not stably associate with light-adapted membranes in either wild-type or Grk1-/- retinas under our experimental conditions, but redistributed to the cone outer segments in a light-dependent manner. The redistribution of transducin subunits to the inner segments in light occurred in both wild-type and Grk1-/-/Arr-/- double knockout photoreceptors. However, Tbetagamma subunits did not redistribute in the absence of Talpha, suggesting that transducin only translocates as an intact heterotrimer.
We conclude that in rods, Arr redistribution requires neither rhodopsin phosphorylation nor phototransduction, suggesting the presence of another light-dependent pathway to trigger translocation. In cones, the light-dependent movement of cArr appears to be independent of stable association with the cone pigments. The light-dependent translocations of Arr and transducin subunits in opposite directions appear to be based on independent mechanisms.
光感受器细胞中光转导成分的光依赖性重新分布在光适应过程中起作用。光照时，视杆和视锥抑制蛋白（Arr和cArr）从内段转运至外段，而转导蛋白亚基（Tα、Tβγ）则向相反方向转运。其潜在的转运机制尚不清楚。本研究在光转导缺陷的小鼠中研究这些先前已证实的转运情况。
在暗适应和光适应的情况下，使用免疫印迹和免疫细胞化学方法检测缺乏G蛋白偶联受体激酶1（Grk1-/-）的单基因敲除小鼠、缺乏GRK1和转导蛋白α亚基（Grk1-/-/Gnat1-/-）或缺乏GRK1和抑制蛋白（Grk1-/-/Arr-/-）的双基因敲除小鼠中Arr、cArr、Tα和Tβγ的分布。
在Grk1-/-小鼠以及Grk1-/-/Gnat1-/-双基因敲除视网膜中，Arr在光照下重新分布至外段。免疫印迹显示，在光适应的野生型、Grk1-/-和Gnat1-/-/Grk1-/-小鼠视网膜中，约25%-50%的Arr与膜结合。相比之下，在我们的实验条件下，无论是野生型还是Grk1-/-视网膜中，cArr在光适应的膜上都不稳定结合，但以光依赖的方式重新分布至视锥外段。在野生型和Grk1-/-/Arr-/-双基因敲除光感受器中，转导蛋白亚基在光照下向内段的重新分布均发生。然而，在没有Tα的情况下，Tβγ亚基不会重新分布，这表明转导蛋白仅作为完整的异源三聚体进行转运。
我们得出结论，在视杆细胞中，Arr的重新分布既不需要视紫红质磷酸化也不需要光转导，这表明存在另一种光依赖途径来触发转运。在视锥细胞中，cArr的光依赖运动似乎独立于与视锥色素的稳定结合。Arr和转导蛋白亚基在相反方向的光依赖转运似乎基于独立的机制。
