Section on Molecular Neuroscience, National Institute of Mental Health Intramural Research Program, Bethesda, MD 20892.
Section on Directed Gene Transfer Laboratory of Cellular and Molecular Regulation, National Institute of Mental Health Intramural Research Program, Bethesda, MD 20892.
eNeuro. 2017 Sep 25;4(5). doi: 10.1523/ENEURO.0248-17.2017. eCollection 2017 Sep-Oct.
The neuritogenic cAMP sensor (NCS), encoded by the gene, links cAMP elevation to activation of extracellular signal-regulated kinase (ERK) in neurons and neuroendocrine cells. Transducing human embryonic kidney (HEK)293 cells, which do not express Rapgef2 protein or respond to cAMP with ERK phosphorylation, with a vector encoding a cDNA reconstituted cAMP-dependent ERK activation. Mutation of a single residue in the cyclic nucleotide-binding domain (CNBD) conserved across cAMP-binding proteins abrogated cAMP-ERK coupling, while deletion of the CNBD altogether resulted in constitutive ERK activation. Two types of mRNA are transcribed from . Rapgef2 protein expression was limited to tissues, i.e., neuronal and endocrine, expressing the second type of mRNA, initiated exclusively from an alternative first exon called here exon 1', and an alternative 5' protein sequence leader fused to a common remaining open reading frame, which is termed here NCS-Rapgef2. In the male mouse brain, NCS-Rapgef2 is prominently expressed in corticolimbic excitatory neurons, and striatal medium spiny neurons (MSNs). Rapgef2-dependent ERK activation by the dopamine D1 agonist SKF81297 occurred in neuroendocrine neuroscreen-1 (NS-1) cells expressing the human D1 receptor and was abolished by deletion of . Corticolimbic [e.g., dentate gyrus (DG), basolateral amygdala (BLA)] ERK phosphorylation induced by SKF81297 was significantly attenuated in ; male mice. ERK phosphorylation in nucleus accumbens (NAc) MSNs induced by treatment with SKF81297, or the psychostimulants cocaine or amphetamine, was abolished in male mice with NAc NCS-Rapgef2-depleting AAV-Synapsin-Cre injections. We conclude that D1-dependent ERK phosphorylation in mouse brain requires NCS-Rapgef2 expression.
神经元生成 cAMP 感受器 (NCS) 由 基因编码，将 cAMP 水平升高与神经元和神经内分泌细胞中细胞外信号调节激酶 (ERK) 的激活联系起来。转染不表达 Rapgef2 蛋白或不通过 cAMP 磷酸化 ERK 的人胚肾 (HEK)293 细胞，用编码 cDNA 的载体重建 cAMP 依赖性 ERK 激活。在跨环核苷酸结合蛋白保守的环核苷酸结合域 (CNBD) 中突变单个残基，阻断了 cAMP-ERK 偶联，而完全删除 CNBD 则导致 ERK 持续激活。从 转录两种类型的 mRNA。Rapgef2 蛋白表达仅限于表达第二种 mRNA 的组织，即神经元和内分泌组织，该 mRNA 仅从称为外显子 1'的替代第一个外显子开始，并且与共同剩余开放阅读框融合的替代 5'蛋白序列先导，在这里称为 NCS-Rapgef2。在雄性小鼠大脑中，NCS-Rapgef2 在皮质边缘兴奋性神经元和纹状体中型棘突神经元 (MSNs) 中表达明显。多巴胺 D1 激动剂 SKF81297 引起的 Rapgef2 依赖性 ERK 激活发生在表达人 D1 受体的神经内分泌神经筛选-1 (NS-1) 细胞中，并通过 删除 而被废除。SKF81297 诱导的皮质边缘 [例如，齿状回 (DG)、基底外侧杏仁核 (BLA)] ERK 磷酸化在 中显著减弱；雄性小鼠。SKF81297 或兴奋剂可卡因或安非他命处理诱导的伏隔核 (NAc) MSNs 中的 ERK 磷酸化在 NAc NCS-Rapgef2 耗竭 AAV-Synapsin-Cre 注射的雄性 小鼠中被消除。我们得出结论，小鼠大脑中 D1 依赖性 ERK 磷酸化需要 NCS-Rapgef2 表达。
