Kanoh J, Watanabe Y, Ohsugi M, Iino Y, Yamamoto M
Department of Biophysics and Biochemistry, School of Science, University of Tokyo, Hongo, Japan.
Genes Cells. 1996 Apr;1(4):391-408. doi: 10.1046/j.1365-2443.1996.d01-247.x.
Fission yeast cells arrest at G1 phase when starved of nitrogen. The molecular mechanism that ensures this arrest is poorly understood. We took a genetic approach to this problem.
The fission yeast gad7-1 mutant failed to arrest at G1 when starved of nitrogen, and was poor in mating and sporulation. The gad7 gene was cloned by complementation. The deduced gad7 gene product was a bZIP protein of 566 amino acids, which could bind to the CRE (cAMP response element) sequence in vitro. Disruption of gad7 resulted in the same phenotypes as gad7-1. Expression of ste11, which encodes a key transcription factor for sexual development, was not inducible in the disruptant. Gad7 was co-immunoprecipitated with another bZIP protein Pcr1, suggesting that the two proteins form a heterodimer in vivo. Gad7 was phosphorylated, and the state of its phosphorylation appeared to be modified in pka1delta or wis1delta cells.
Gad7, a CRE-binding protein that cooperates with Pcr1, is required for proper G1 arrest and gene expression under nitrogen starvation. Gad7 is a phosphoprotein, whose activity may be regulated by protein kinases including the cAMP-dependent protein kinase (Pka1) and Wis1 osmosensory MAP kinase kinase.
裂殖酵母细胞在氮饥饿时会停滞在G1期。但确保这种停滞的分子机制仍知之甚少。我们采用遗传学方法来研究这个问题。
裂殖酵母gad7-1突变体在氮饥饿时无法停滞在G1期，且交配和孢子形成能力较差。通过互补作用克隆了gad7基因。推导的gad7基因产物是一个由566个氨基酸组成的bZIP蛋白，它在体外能与CRE（cAMP反应元件）序列结合。gad7的破坏导致了与gad7-1相同的表型。编码性发育关键转录因子的ste11在破坏突变体中无法被诱导表达。Gad7与另一个bZIP蛋白Pcr1共同免疫沉淀，表明这两种蛋白在体内形成异源二聚体。Gad7被磷酸化，其磷酸化状态在pka1δ或wis1δ细胞中似乎发生了改变。
Gad7是一种与Pcr1合作的CRE结合蛋白，在氮饥饿条件下对于正确的G1期停滞和基因表达是必需的。Gad7是一种磷蛋白，其活性可能受包括cAMP依赖性蛋白激酶（Pka1）和Wis1渗透压感应丝裂原活化蛋白激酶激酶在内的蛋白激酶调节。
