Huber A, Belusic G, Da Silva N, Bähner M, Gerdon G, Draslar K, Paulsen R
Institute of Zoology, Department of Cell Biology and Neurobiology, University of Karlsruhe, Haid-und-Neu-Str. 9, D-76131 Karlsruhe, Germany.
Eur J Neurosci. 2000 Nov;12(11):3909-18. doi: 10.1046/j.1460-9568.2000.00276.x.
The visual transduction cascade of fly photoreceptors is a G protein-coupled phospholipase C-signalling pathway which is assembled into a supramolecular signalling complex by the PDZ (postsynaptic density protein-95, discs large, Z0-1) domain protein INAD (inactivation no afterpotential D). The norpA-encoded phospholipase Cbeta, the light-activated transient receptor potential (TRP) Ca2+ channel and an eye-specific protein kinase C are bound to INAD and together form the core of the signalling complex. In the present study we show that the Calliphora rpa mutant, which has previously been hypothesized to represent an equivalent of Drosophila norpA mutants, has normal amounts of norpA mRNA but fails to express inaD mRNA. Electrophysiological recordings from the eyes of the rpa mutant reveal that the electroretinogram is reduced (about 12% of wild type) but not completely absent, and that it exhibits markedly prolonged deactivation kinetics. Furthermore, rpa mutants display a slow, light-dependent degeneration of the photoreceptor cells. With respect to the INAD signalling complex, the rpa mutant is similar to the Drosophila inaD null mutant: not only INAD itself, but also the other core components of the INAD signalling complex, are reduced or absent in photoreceptor membranes of rpa flies. Residual TRP is localized throughout the plasma membrane of the photoreceptor cell, rather than being restricted to the microvillar photoreceptor membrane. [35S]methionine-labelling of newly synthesized retinal proteins reveals that TRP is synthesized in the rpa mutant at wild-type level, but is transported to or incorporated into the microvillar photoreceptor membrane at a much lower rate. We thus suggest, that the formation of the INAD signalling complex is required for specifically targeting its components to the photoreceptor membrane.
果蝇光感受器的视觉转导级联是一种G蛋白偶联的磷脂酶C信号通路，它通过PDZ（突触后致密蛋白95、盘大蛋白、Z0 - 1）结构域蛋白INAD（无后电位失活D）组装成超分子信号复合物。由norpA编码的磷脂酶Cβ、光激活的瞬时受体电位（TRP）Ca²⁺通道和一种眼特异性蛋白激酶C与INAD结合，共同构成信号复合物的核心。在本研究中，我们发现丽蝇rpa突变体，此前被假设相当于果蝇norpA突变体，其norpA mRNA含量正常，但未能表达inaD mRNA。对rpa突变体眼睛的电生理记录显示，视网膜电图降低（约为野生型的12%）但并非完全消失，并且其失活动力学明显延长。此外，rpa突变体表现出光感受器细胞缓慢的、光依赖性退化。关于INAD信号复合物，rpa突变体类似于果蝇inaD基因敲除突变体：不仅INAD本身，而且INAD信号复合物的其他核心成分，在rpa果蝇的光感受器膜中减少或缺失。残余的TRP定位于光感受器细胞的整个质膜，而不是局限于微绒毛光感受器膜。用[³⁵S]甲硫氨酸标记新合成的视网膜蛋白表明，TRP在rpa突变体中以野生型水平合成，但以低得多的速率转运至或整合到微绒毛光感受器膜中。因此，我们认为，INAD信号复合物的形成是将其组分特异性靶向到光感受器膜所必需的。
