Schadick Kevin, Fourcade H Matthew, Boumenot Peter, Seitz Jeffrey J, Morrell Jennifer L, Chang Louise, Gould Kathleen L, Partridge Janet F, Allshire Robin C, Kitagawa Katsumi, Hieter Phil, Hoffman Charles S
Biology Department, Boston College, Chestnut Hill Massachusetts 02467, USA.
Eukaryot Cell. 2002 Aug;1(4):558-67. doi: 10.1128/EC.1.4.558-567.2002.
The Schizosaccharomyces pombe fbp1 gene, encoding fructose-1,6-bisphosphatase, is transcriptionally repressed by glucose. Mutations that confer constitutive fbp1 transcription identify git (glucose-insensitive transcription) genes that encode components of a cyclic AMP (cAMP) signaling pathway required for adenylate cyclase activation. Four of these genes encode the three subunits of a heterotrimeric G protein (gpa2, git5, and git11) and a G protein-coupled receptor (git3). Three additional genes, git1, git7, and git10, act in parallel to or downstream from the G protein genes. Here, we describe the cloning and characterization of the git7 gene. The Git7p protein is a member of the Saccharomyces cerevisiae Sgtlp protein family. In budding yeast, Sgtlp associates with Skplp and plays an essential role in kinetochore assembly, while in Arabidopsis, a pair of SGT1 proteins have been found to be involved in plant disease resistance through an interaction with RAR1. Like S. cerevisiae Sgtlp, Git7p is essential, but this requirement appears to be due to roles in septation and cell wall integrity, which are unrelated to cAMP signaling, as S. pombe cells lacking either adenylate cyclase or protein kinase A are viable. In addition, git7 mutants are sensitive to the microtubule-destabilizing drug benomyl, although they do not display a chromosome stability defect. Two alleles of git7 that are functional for cell growth and septation but defective for glucose-triggered cAMP signaling encode proteins that are altered in the highly conserved carboxy terminus. The S. cerevisiae and human SGT1 genes both suppress git7-93 but not git7-235 for glucose repression of fbp1 transcription and benomyl sensitivity. This allele-specific suppression indicates that the Git7p/Sgtlp proteins may act as multimers, such that Git7-93p but not Git7-235p can deliver the orthologous proteins to species-specific targets. Our studies suggest that members of the Git7p/Sgt1p protein family may play a conserved role in the regulation of adenylate cyclase activation in S. pombe, S. cerevisiae, and humans.
粟酒裂殖酵母fbp1基因编码果糖-1,6-二磷酸酶，其转录受葡萄糖抑制。赋予fbp1组成型转录的突变可鉴定出git（葡萄糖不敏感转录）基因，这些基因编码激活腺苷酸环化酶所需的环磷酸腺苷（cAMP）信号通路的组分。其中四个基因编码异源三聚体G蛋白的三个亚基（gpa2、git5和git11）以及一个G蛋白偶联受体（git3）。另外三个基因git1、git7和git10与G蛋白基因平行作用或在其下游起作用。在此，我们描述了git7基因的克隆和特性分析。Git7p蛋白是酿酒酵母Sgtlp蛋白家族的成员。在芽殖酵母中，Sgtlp与Skplp相关联，并在动粒组装中起关键作用，而在拟南芥中，已发现一对SGT1蛋白通过与RAR1相互作用参与植物抗病性。与酿酒酵母Sgtlp一样，Git7p是必需的，但这种需求似乎是由于其在隔膜形成和细胞壁完整性方面的作用，这与cAMP信号传导无关，因为缺乏腺苷酸环化酶或蛋白激酶A的粟酒裂殖酵母细胞是有活力的。此外，git7突变体对微管解聚药物苯菌灵敏感，尽管它们没有表现出染色体稳定性缺陷。两个对细胞生长和隔膜形成有功能但对葡萄糖触发的cAMP信号传导有缺陷的git7等位基因编码在高度保守的羧基末端发生改变的蛋白质。酿酒酵母和人类SGT1基因在fbp1转录的葡萄糖抑制和苯菌灵敏感性方面均能抑制git7-93而不能抑制git7-235。这种等位基因特异性抑制表明Git7p/Sgtlp蛋白可能作为多聚体起作用，使得Git7-93p而非Git7-235p能够将直系同源蛋白传递到物种特异性靶点。我们的研究表明，Git7p/Sgt1p蛋白家族成员可能在粟酒裂殖酵母、酿酒酵母和人类中腺苷酸环化酶激活的调节中起保守作用。
