Molecular Medicine Program, Taiwan International Graduate Program, Academia Sinica, Taipei 115, Taiwan.
Biochem J. 2010 Dec 1;432(2):267-73. doi: 10.1042/BJ20100617.
GC (guanylate cyclase)-G is the most recently identified member of the receptor GC family. However, the regulation of its activity and protein expression in the mammalian olfactory system remains unclear. In the present study, we used a GC-G-specific antibody to validate that the GC-G protein is expressed in Grueneberg ganglion neurons, a newly recognized olfactory subsystem co-expressing other cGMP signalling components such as the cGMP-regulated PDE2A (phosphodiesterase 2A) and the cGMP-gated ion channel CNGA3 (cyclic nucleotide-gated cation channel α-3). Further molecular and biochemical analyses showed that heterologously expressed GC-G protein, specifically the C-terminal cyclase domain, was directly stimulated by bicarbonate in both in vivo cellular cGMP accumulation assays in human embryonic kidney-293T cells and in vitro GC assays with a purified recombinant protein containing the GC domain. In addition, overexpression of GC-G in NG108 neuronal cells resulted in a CO2-dependent increase in cellular cGMP level that could be blocked by treatment with acetazolamide, an inhibitor of carbonic anhydrases, which implies that the stimulatory effect of CO2 requires its conversion to bicarbonate. Together, our data demonstrate a novel CO2/bicarbonate-dependent activation mechanism for GC-G and suggest that GC-G may be involved in a wide variety of CO2/bicarbonate-regulated biological processes such as the chemosensory function in Grueneberg ganglion neurons.
GC（鸟苷酸环化酶）-G 是最近被发现的受体 GC 家族成员。然而，其在哺乳动物嗅觉系统中的活性和蛋白表达的调控仍不清楚。在本研究中，我们使用 GC-G 特异性抗体验证了 GC-G 蛋白在 Grueneberg 神经节神经元中表达，该神经元是一个新发现的嗅觉子系统，共同表达其他 cGMP 信号成分，如 cGMP 调节的 PDE2A（磷酸二酯酶 2A）和 cGMP 门控离子通道 CNGA3（环核苷酸门控阳离子通道 α-3）。进一步的分子和生化分析表明，异源表达的 GC-G 蛋白，特别是其 C 端环化酶结构域，可被碳酸氢盐直接刺激，这在人胚肾 293T 细胞中的体内细胞 cGMP 积累测定和含有 GC 结构域的纯化重组蛋白的体外 GC 测定中均得到证实。此外，在 NG108 神经元细胞中过表达 GC-G 导致细胞 cGMP 水平的 CO2 依赖性增加，该增加可被碳酸酐酶抑制剂乙酰唑胺所阻断，这表明 CO2 的刺激作用需要其转化为碳酸氢盐。总之，我们的数据证明了 GC-G 的一种新型 CO2/碳酸氢盐依赖性激活机制，并提示 GC-G 可能参与各种 CO2/碳酸氢盐调节的生物学过程，如 Grueneberg 神经节神经元的化学感觉功能。
