Identification and characterization of AGS4 -: A protein containing three G-protein regulatory motifs that regulate the activation state of Giα
Identification and characterization of AGS4 -: A protein containing three G-protein regulatory motifs that regulate the activation state of Giα
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DOI：
10.1074/jbc.m312786200
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发表时间：
2004-06-25
影响因子：
4.8
通讯作者：
Lanier, SM
中科院分区：
文献类型：
--
作者：
Cao, XQ;Cismowski, MJ;Lanier, SM
在哺乳动物 cDNA 的功能筛选中鉴定出 G 蛋白信号传导 1-3 (AGS1-3) 激活剂，该 cDNA 在没有受体的情况下激活 G 蛋白信号传导。我们报告了从人前列腺平滑肌肉瘤 cDNA 文库中分离和表征了另一种 AGS 蛋白 (AGS4)。 AGS4 与 G18.1b 相同，G18.1b 由 6 号染色体主要组织相容性 III 类区域内的基因编码。基于酵母的功能筛选中 AGS4 的活性对 G(i2)/G(i3) 具有选择性，并且与 G(i)α 的鸟嘌呤核苷酸交换无关。 RNA 印迹表明 AGS4/G18.1b mRNA 在心脏、胎盘、肺和肝脏中富集。 AGS4/G18.1b 特异性抗血清的免疫细胞化学表明，在 COS7 或中国仓鼠卵巢细胞中表达后，细胞内主要存在非均质、核外分布。 AGS4/G18.1b 在具有多个脯氨酸的氨基末端结构域下游包含三个 G 蛋白调节基序。谷胱甘肽 S-转移酶 (GST)-AGS4/G18.1b 融合蛋白与纯化的 G(i)alpha 相互作用，并且源自每个 G 蛋白调节基序的肽抑制鸟苷 5'-3-O-(硫代)三磷酸 (GTPgammaS) 与纯化的 G(i)alpha(1) 结合。 AGS4/G18.1b 还与细胞转染后 COS7 细胞裂解物中的 G(i)alpha(3) 复合。然而，AGS4/G18.1b 并没有改变与 Gbetagamma 调节的效应磷脂酶 C-beta2 共转染的 COS7 细胞中肌醇磷酸盐的产生。这些数据表明，要么需要额外的信号将AGS4/G18.1b定位在正确的细胞位置，在那里它可以接触异源三聚体并促进亚基解离，要么AGS4作为G(i)alpha的替代结合伴侣，独立于Gbetagamma参与G蛋白信号转导事件，而G蛋白信号转导事件独立于细胞表面的经典G蛋白偶联受体。
Activators of G-protein signaling 1-3 (AGS1-3) were identified in a functional screen of mammalian cDNAs that activated G-protein signaling in the absence of a receptor. We report the isolation and characterization of an additional AGS protein (AGS4) from a human prostate leiomyosarcoma cDNA library. AGS4 is identical to G18.1b, which is encoded by a gene within the major histocompatibility class III region of chromosome 6. The activity of AGS4 in the yeast-based functional screen was selective for G(i2)/G(i3) and independent of guanine-nucleotide exchange by G(i)alpha. RNA blots indicated enrichment of AGS4/G18.1b mRNA in heart, placenta, lung, and liver. Immunocytochemistry with AGS4/G18.1b-specific antisera indicated a predominant nonhomogeneous, extranuclear distribution within the cell following expression in COS7 or Chinese hamster ovary cells. AGS4/G18.1b contains three G-protein regulatory motifs downstream of an amino terminus domain with multiple prolines. Glutathione S-transferase (GST)-AGS4/G18.1b fusion proteins interacted with purified G(i)alpha, and peptides derived from each of the G-protein regulatory motifs inhibited guanosine 5'-3-O-(thio)triphosphate (GTPgammaS) binding to purified G(i)alpha(1). AGS4/G18.1b was also complexed with G(i)alpha(3) in COS7 cell lysates following cell transfection. However, AGS4/G18.1b did not alter the generation of inositol phosphates in COS7 cells cotransfected with the Gbetagamma-regulated effector phospholipase C-beta2. These data suggest either that an additional signal is required to position AGS4/G18.1b in the proper cellular location where it can access heterotrimer and promote subunit dissociation or that AGS4 serves as an alternative binding partner for G(i)alpha independent of Gbetagamma participating in G-protein signaling events that are independent of classical G-protein-coupled receptors at the cell surface.