Buller A L, White M M
Department of Pharmacology, University of Pennsylvania School of Medicine, Philadelphia 19104.
Proc Natl Acad Sci U S A. 1988 Nov;85(22):8717-21. doi: 10.1073/pnas.85.22.8717.
The nicotinic acetylcholine receptor (AcChoR) of Torpedo electroplax is a multisubunit transmembrane glycoprotein complex with a subunit stoichiometry of alpha 2 beta gamma delta. The RNAs for the separate subunits were transcribed in vitro from cDNAs inserted in pSP64T vectors and microinjected in Xenopus oocytes. Microinjected in vitro-transcribed RNAs were stable, with a half-life of 72 hr. Xenopus oocytes assembled functional AcChoRs from the subunit-specific RNAs. These receptors were inserted in the cell membrane and could be detected as early as 6 hr after RNA microinjection. The oocyte-expressed AcChoR subunits could be immunoprecipitated with anti-Torpedo AcChoR subunit antibodies. Expression of the AcChoR in oocytes proceeded linearly for 72 hr after microinjection. While the amount of RNA injected did not alter the linearity of the expression time course, the rate of receptor expression in oocytes showed a saturable dependence on RNA concentration. Varying the relative amount of alpha-subunit RNA microinjected into oocytes had a striking effect on receptor expression. This effect was specific for the alpha-subunit. These results suggest that transcript availability may control receptor expression in Xenopus oocytes. In addition, the availability of the alpha-subunit may be a limiting factor for receptor expression.
电鳐电器官的烟碱型乙酰胆碱受体（AcChoR）是一种多亚基跨膜糖蛋白复合体，其亚基化学计量比为α2βγδ。各个亚基的RNA是从插入pSP64T载体的cDNA体外转录而来，并显微注射到非洲爪蟾卵母细胞中。显微注射的体外转录RNA是稳定的，半衰期为72小时。非洲爪蟾卵母细胞利用亚基特异性RNA组装功能性AcChoRs。这些受体插入细胞膜，早在RNA显微注射后6小时就能被检测到。卵母细胞表达的AcChoR亚基可用抗电鳐AcChoR亚基抗体进行免疫沉淀。显微注射后，卵母细胞中AcChoR的表达在72小时内呈线性进行。虽然注射的RNA量不会改变表达时间进程的线性，但卵母细胞中受体的表达速率对RNA浓度呈现出饱和依赖性。改变显微注射到卵母细胞中的α亚基RNA的相对量对受体表达有显著影响。这种影响对α亚基具有特异性。这些结果表明，转录本的可利用性可能控制非洲爪蟾卵母细胞中的受体表达。此外，α亚基的可利用性可能是受体表达的一个限制因素。
