Welton R M, Hoffman C S
Department of Biology, Boston College, Chestnut Hill, Massachusetts 02467, USA.
Genetics. 2000 Oct;156(2):513-21. doi: 10.1093/genetics/156.2.513.
The fission yeast Schizosaccharomyces pombe responds to environmental glucose by activating adenylate cyclase. The resulting cAMP signal activates protein kinase A (PKA). PKA inhibits glucose starvation-induced processes, such as conjugation and meiosis, and the transcription of the fbp1 gene that encodes the gluconeogenic enzyme fructose-1,6-bisphosphatase. We previously identified a collection of git genes required for glucose repression of fbp1 transcription, including pka1/git6, encoding the PKA catalytic subunit, git2/cyr1, encoding adenylate cyclase, and six "upstream" genes required for adenylate cyclase activation. The git8 gene, identical to gpa2, encodes the alpha subunit of a heterotrimeric guanine-nucleotide binding protein (Galpha) while git5 encodes a Gbeta subunit. Multicopy suppression studies with gpa2(+) previously indicated that S. pombe adenylate cyclase activation may resemble that of the mammalian type II enzyme with sequential activation by Galpha followed by Gbetagamma. We show here that an activated allele of gpa2 (gpa2(R176H), carrying a mutation in the coding region for the GTPase domain) fully suppresses mutations in git3 and git5, leading to a refinement in our model. We describe the cloning of git3 and show that it encodes a putative seven-transmembrane G protein-coupled receptor. A git3 deletion confers the same phenotypes as deletions of other components of the PKA pathway, including a germination delay, constitutive fbp1 transcription, and starvation-independent conjugation. Since the git3 deletion is fully suppressed by the gpa2(R176H) allele with respect to fbp1 transcription, git3 appears to encode a G protein-coupled glucose receptor responsible for adenylate cyclase activation in S. pombe.
裂殖酵母粟酒裂殖酵母通过激活腺苷酸环化酶对环境中的葡萄糖作出反应。产生的cAMP信号激活蛋白激酶A（PKA）。PKA抑制葡萄糖饥饿诱导的过程，如接合和减数分裂，以及编码糖异生酶果糖-1,6-二磷酸酶的fbp1基因的转录。我们之前鉴定了一组fbp1转录的葡萄糖抑制所需的git基因，包括编码PKA催化亚基的pka1/git6、编码腺苷酸环化酶的git2/cyr1，以及腺苷酸环化酶激活所需的六个“上游”基因。与gpa2相同的git8基因编码异源三聚体鸟嘌呤核苷酸结合蛋白（Gα）的α亚基，而git5编码Gβ亚基。之前用gpa2(+)进行的多拷贝抑制研究表明，粟酒裂殖酵母腺苷酸环化酶的激活可能类似于哺乳动物II型酶，由Gα随后是Gβγ顺序激活。我们在此表明，gpa2的一个激活等位基因（gpa2(R176H)，在GTPase结构域的编码区域携带一个突变）完全抑制git3和git5中的突变，从而使我们的模型得到完善。我们描述了git3的克隆，并表明它编码一个推定的七跨膜G蛋白偶联受体。git3缺失赋予与PKA途径其他组分缺失相同的表型，包括萌发延迟、fbp1的组成型转录以及与饥饿无关的接合。由于就fbp1转录而言，git3缺失被gpa2(R176H)等位基因完全抑制，git3似乎编码一个负责粟酒裂殖酵母中腺苷酸环化酶激活的G蛋白偶联葡萄糖受体。
