Stearns T, Kahn R A, Botstein D, Hoyt M A
Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
Mol Cell Biol. 1990 Dec;10(12):6690-9. doi: 10.1128/mcb.10.12.6690-6699.1990.
ADP ribosylation factor (ARF) is a ubiquitous 21-kDa GTP-binding protein in eucaryotes. ARF was first identified in animal cells as the protein factor required for the efficient ADP-ribosylation of the mammalian G protein Gs by cholera toxin in vitro. A gene (ARF1) encoding a protein homologous to mammalian ARF was recently cloned from Saccharomyces cerevisiae (Sewell and Kahn, Proc. Natl. Acad. Sci. USA, 85:4620-4624, 1988). We have found a second gene encoding ARF in S. cerevisiae, ARF2. The two ARF genes are within 28 centimorgans of each other on chromosome IV, and the proteins encoded by them are 96% identical. Disruption of ARF1 causes slow growth, cold sensitivity, and sensitivity to normally sublethal concentrations of fluoride ion in the medium. Disruption of ARF2 causes no detectable phenotype. Disruption of both genes is lethal; thus, ARF is essential for mitotic growth. The ARF1 and ARF2 proteins are functionally homologous, and the phenotypic differences between mutations in the two genes can be accounted for by the level of expression; ARF1 produces approximately 90% of total ARF. Among revertants of the fluoride sensitivity of an arf1 null mutation were ARF1-ARF2 fusion genes created by a gene conversion event in which the deleted ARF1 sequences were repaired by recombination with ARF2.
ADP核糖基化因子(ARF)是真核生物中普遍存在的一种21kDa的GTP结合蛋白。ARF最初是在动物细胞中被鉴定出来的,它是霍乱毒素在体外高效催化哺乳动物G蛋白Gs进行ADP核糖基化所需的蛋白质因子。最近,从酿酒酵母中克隆出了一个编码与哺乳动物ARF同源蛋白的基因(ARF1)(Sewell和Kahn,《美国国家科学院院刊》,85:4620 - 4624,1988)。我们在酿酒酵母中发现了第二个编码ARF的基因,即ARF2。这两个ARF基因在第四条染色体上彼此相距28厘摩,它们编码的蛋白质有96%的同一性。ARF1基因的破坏会导致生长缓慢、对冷敏感以及对培养基中正常情况下亚致死浓度的氟离子敏感。ARF2基因的破坏未导致可检测到的表型。两个基因都被破坏是致死的;因此,ARF对于有丝分裂生长是必不可少的。ARF1和ARF2蛋白在功能上是同源的,两个基因中突变之间的表型差异可以通过表达水平来解释;ARF1产生的ARF约占总量的90%。在arf1缺失突变体对氟敏感性的回复突变体中,有通过基因转换事件产生的ARF1 - ARF2融合基因,在该事件中,缺失的ARF1序列通过与ARF2重组得以修复。