非洲爪蟾卵母细胞腺苷酸环化酶的分子克隆与表达

Molecular cloning and expression of an adenylyl cyclase from Xenopus laevis oocytes.

作者信息

Torrejón M, Echeverría V, Retamales G, Herrera L, Hinrichs M V, Olate J

机构信息

Departamento de Fisiopatología, Facultad de Ciencias Biológicas, Universidad de Concepción, Chile.

出版信息

FEBS Lett. 1997 Mar 3;404(1):91-4. doi: 10.1016/s0014-5793(97)00100-2.

DOI:10.1016/s0014-5793(97)00100-2

PMID:9074644

Abstract

We have cloned a cDNA that encodes a novel Xenopus laevis oocyte adenylyl cyclase (xlAC) using oligonucleotides against conserved mammalian adenylyl cyclase regions. The isolated cDNA is 4372 bp long with an open reading frame of 4065 nucleotides which encodes a protein of 1355 amino acids. Comparison of the deduced amino acid sequence with previously cloned mammalian adenylyl cyclases shows a low identity, 19.7% with type 2 rat adenylyl cyclase and 24.2% with type 4 rat adenylyl cyclase, indicating that this Xenopus isoform represents a new member of this protein family. Gene expression studies of the xlAC by reverse PCR showed that this gene is expressed in all oogenesis stages but not during early embryogenesis. Expression of the xlAC in COS-7 cells resulted in increased basal AC activity, that was stimulated by forskolin, Gpp(NH)p and aluminium fluoride, and was insensitive to calcium and calcium-calmodulin (Ca2(+)-CaM).

摘要

我们利用针对保守的哺乳动物腺苷酸环化酶区域的寡核苷酸，克隆了一个编码新型非洲爪蟾卵母细胞腺苷酸环化酶（xlAC）的cDNA。分离得到的cDNA长4372 bp，具有一个4065个核苷酸的开放阅读框，编码一个1355个氨基酸的蛋白质。将推导的氨基酸序列与先前克隆的哺乳动物腺苷酸环化酶进行比较，发现同源性较低，与大鼠2型腺苷酸环化酶的同源性为19.7%，与大鼠4型腺苷酸环化酶的同源性为24.2%，这表明这种非洲爪蟾同工型代表了该蛋白家族的一个新成员。通过逆转录PCR对xlAC进行基因表达研究表明，该基因在卵母细胞发生的所有阶段均有表达，但在早期胚胎发育过程中不表达。xlAC在COS-7细胞中的表达导致基础AC活性增加，该活性受到福斯可林、Gpp(NH)p和氟化铝的刺激，且对钙和钙调蛋白（Ca2(+)-CaM）不敏感。