Grammatopoulos D K, Randeva H S, Levine M A, Katsanou E S, Hillhouse E W
Sir Quinton Hazell Molecular Medicine Research Centre, Department of Biological Sciences, University of Warwick, Coventry, United Kingdom.
Mol Endocrinol. 2000 Dec;14(12):2076-91. doi: 10.1210/mend.14.12.0574.
CRH and CRH-related peptides such as urocortin mediate their actions in the human myometrium via activation of two distinct classes of CRH receptors, R1 and R2. These heptahelical receptors are able to stimulate a number of different intracellular signals; one key mediator of G protein-activated intracellular signaling is the cascade of p42/p44, mitogen-activated protein kinase (MAPK). We therefore hypothesized that activation of MAPK might mediate CRH and or/urocortin actions in the myometrium. In cultured human pregnant myometrial cells, urocortin but not CRH was able to induce MAPK phosphorylation and activation, suggesting that in the human myometrium these two peptides have distinct actions and biological roles. To identify the particular receptor subtypes mediating this phenomenon, all known CRH receptors present in the human myometrial cells were stably expressed individually in HEK293 and CHO cells, and their ability to activate MAPK was tested. The R1alpha and R2beta, but not the R1beta, R1c, or R1d, receptor subtypes were able to mediate urocortin-induced MAPK activation. The signaling components were further investigated; activation of Gs, Go, or Gi proteins did not appear to be involved, but activation of Gq with subsequent production of inositol triphosphates (IP3) and protein kinase C (PKC) activation correlated with MAPK phosphorylation. Studies on Gq protein activation using [alpha-32P]-GTP-gamma-azidoanilide and IP3 production in cells expressing the R1alpha or R2beta CRH receptors demonstrated that urocortin was 10 times more potent than CRH. Moreover, urocortin (UCN) generated peak responses that were 50-70% greater than CRH in activating the Gq protein and stimulating IP3 production. In conclusion, UCN acting thought multiple receptor subtypes can stimulate myometrial MAPK via induction of the Gq/phospholipase C/IP3/PKC pathway, whereas CRH-induced activation of this pathway appears to be insufficient to achieve MAPK activation.
促肾上腺皮质激素释放激素（CRH）以及诸如尿皮质素等与CRH相关的肽，通过激活两类不同的CRH受体R1和R2，在人子宫肌层中发挥作用。这些七螺旋受体能够刺激多种不同的细胞内信号；G蛋白激活的细胞内信号传导的一个关键介质是p42/p44丝裂原活化蛋白激酶（MAPK）级联反应。因此我们推测，MAPK的激活可能介导CRH和/或尿皮质素在子宫肌层中的作用。在培养的人妊娠子宫肌层细胞中，尿皮质素而非CRH能够诱导MAPK磷酸化和激活，这表明在人子宫肌层中这两种肽具有不同的作用和生物学功能。为了确定介导这一现象的特定受体亚型，将人子宫肌层细胞中存在的所有已知CRH受体分别在HEK293和CHO细胞中稳定表达，并测试它们激活MAPK的能力。R1α和R2β受体亚型能够介导尿皮质素诱导的MAPK激活，而R1β、R1c或R1d受体亚型则不能。对信号传导成分进行了进一步研究；似乎不涉及Gs、Go或Gi蛋白的激活，但Gq的激活以及随后三磷酸肌醇（IP3）的产生和蛋白激酶C（PKC）的激活与MAPK磷酸化相关。使用[α-32P]-GTP-γ-叠氮苯胺对表达R1α或R2β CRH受体的细胞进行Gq蛋白激活和IP3产生的研究表明，尿皮质素的效力比CRH高10倍。此外，在激活Gq蛋白和刺激IP3产生方面，尿皮质素（UCN）产生的峰值反应比CRH大50 - 70%。总之，UCN通过多种受体亚型发挥作用，可通过诱导Gq/磷脂酶C/IP3/PKC途径刺激子宫肌层MAPK，而CRH诱导的该途径激活似乎不足以实现MAPK激活。
