Zhang Xue, Wensel Theodore G, Kraft Timothy W
Department of Biochemistry and Molecular Biology, Baylor College of Medicine, Houston, Texas 77030, USA.
J Neurosci. 2003 Feb 15;23(4):1287-97. doi: 10.1523/JNEUROSCI.23-04-01287.2003.
Vertebrate cone and rod photoreceptor cells use similar mechanisms to transduce light signals into electrical signals, but their responses to light differ in sensitivity and kinetics. To assess the role of G-protein GTP hydrolysis kinetics in mammalian cone photoresponses, we have characterized photoresponses and GTPase regulatory components of cones and rods from the cone-dominant retina of the eastern chipmunk. Sensitivity, based on the stimulus strength required for a half-maximum response, of the M-cone population was 38-fold lower than that of the rods. The relatively lower cone sensitivity could be attributed in part to lower amplification in the rising phase and in part to faster recovery kinetics. At a molecular level, cloning of chipmunk cDNA and expression of recombinant proteins provided standards for quantitative immunoblot analysis of proteins involved in GTPase acceleration. The ratio of the cGMP-phosphodiesterase inhibitory subunit gamma to cone pigment, 1:68, was similar to the levels observed for ratios to rhodopsin in bovine retina, 1:76, or mouse retina, 1:65. In contrast, the ratio to pigment of the GTPase-accelerating protein RGS9-1 was 1:62, more than 10 times higher than ratios observed in rod-dominant retinas. Immunoprecipitation experiments revealed that, in contrast to rods, RGS9-1 in chipmunk retina is associated with both the short and long isoforms of its partner subunit G(beta5). The much higher levels of the GTPase-accelerating protein complex in cones, compared with rods, suggest a role for GTPase acceleration in obtaining rapid photoresponse kinetics.
脊椎动物的视锥和视杆光感受器细胞利用相似的机制将光信号转化为电信号，但其对光的反应在敏感性和动力学方面存在差异。为了评估G蛋白GTP水解动力学在哺乳动物视锥光反应中的作用，我们对东部花栗鼠视锥占主导的视网膜中的视锥和视杆的光反应及GTP酶调节成分进行了表征。基于产生半数最大反应所需的刺激强度，M视锥群体的敏感性比视杆低38倍。视锥相对较低的敏感性部分可归因于上升阶段较低的放大倍数，部分可归因于更快的恢复动力学。在分子水平上，花栗鼠cDNA的克隆和重组蛋白的表达为参与GTP酶加速的蛋白质的定量免疫印迹分析提供了标准。环鸟苷酸磷酸二酯酶抑制亚基γ与视锥色素的比例为1:68，与在牛视网膜中与视紫红质的比例（1:76）或小鼠视网膜中与视紫红质的比例（1:65）相似。相比之下，GTP酶加速蛋白RGS9-1与色素的比例为1:62，比在视杆占主导的视网膜中观察到的比例高出10倍以上。免疫沉淀实验表明，与视杆不同，花栗鼠视网膜中的RGS9-1与其伙伴亚基G(β5)的短异构体和长异构体都有关联。与视杆相比，视锥中GTP酶加速蛋白复合物的水平要高得多，这表明GTP酶加速在获得快速光反应动力学中起作用。
