GRP-R AND NMB-R SIGNALLING

使用纯化的异源三聚体GTP结合蛋白的重构研究 脂囊泡中的Gq（G蛋白）表明， 与Gs、Gi和Go亚家族相反， 磷脂酶C-1在配体受体相互作用后， 百日咳毒素 我们假设Gq或结构相似的 Galpha 11在蛙皮素受体信号转导中起重要作用。 我们 通过观察Gq-和Galpha 11-的影响， 蛙皮素特异性反义分子对爪蟾免疫应答的影响 表达GRP-R或NMB-R的卵母细胞。 合理设计反义核酸 寡核苷酸，我们已经分离和测序非洲爪蟾的cDNA克隆 Gq和Galpha 11。 非洲爪蟾的蛋白质在氨基酸上有超过90%的相同性 与哺乳动物对应物的氨基酸序列。 这些序列被用来 设计合成反义硫代磷酸寡核苷酸 将其注射到表达GRP-R或NMB-R的非洲爪蟾卵母细胞中， 观察它们对蛙皮素信号传导的影响。反义 Gq特异性寡核苷酸抑制NMB-R信号传导，但不 影响GRP-R信号传导。 与此相反，Galpha 11反义 寡核苷酸不改变来自GRP-R或NMB-R的信号传导。 这些研究表明，虽然信号转导途径， GRP-R和NMB-R相似，两种受体的G蛋白不同 耦合特性 此外，Gq和Galpha 11不是 可互换用于NMB-R联轴器。 我们假设 Gq家族成员的表达可用于调节受体 不同细胞的反应。

Reconstitution studies using purified heterotrimeric GTP-binding proteins (G-proteins) in lipid vesicles indicate that the Gq sub-family of alpha subunits, in contrast to the Gs, Gi, and Go subfamilies, activate phospholipase C-1 after ligand receptor interaction, and are insensitive to pertussis toxin. We postulate that Gq, or structurally similar Galpha11, are important for bombesin receptor signal transduction. We have tested this hypothesis by observing the effects of Gq- and Galpha11- specific antisense molecules on bombesin-mediated responses in Xenopus oocytes expressing GRP-R or NMB-R. To rationally design antisense oligonucleotides, we have isolated and sequenced cDNA clones for Xenopus Gq and Galpha11. The Xenopus proteins are over 90% identical in amino acid sequence to their mammalian counterparts. These sequences were used to design the synthesis of antisense phosphothiorate oligonucleotides which were injected into Xenopus oocytes expressing either GRP-R or NMB-R to observe their effects on bombesin signalling. The antisense oligonucleotide specific for Gq inhibited NMB-R signalling, but did not effect GRP-R signalling. In contrast, the Galpha11 antisense oligonucleotide did not alter signalling from either GRP-R or NMB-R. These studies indicate that, although the signal transduction pathway for GRP-R and NMB-R is similar, the two receptors differ in their G-protein coupling properties. In addition, Gq and Galpha11 are not interchangeable for NMB-R coupling. We hypothesize that the pattern of expression of Gq family members may be used to modulate receptor responses in different cells.