Studies on the regulation of the expression of circadian clock gene cluster kaiABC by using bioluminescence as a reporter

以生物发光为报告基因的生物钟基因簇kaiABC表达调控研究

• 批准号: 08454244
• 负责人: ISHIURA Masahiro
• 金额: $ 4.8万
• 依托单位: Nagoya University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1997
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08454244/
• 关键词: Circadian clock gene; Bioluminescence; Gene expression; Cyanobacteria; Luciferase; kaiABC; Promoters; プロモーター; サーカディアンリズム; 生物時計; 時計遺伝子; Kaiタンパク質; Synechococcus sp.PCC 7942; Sipeihaiecmis.; オペロン; 生物時計突然変異体

We transferred a genomic DNA library into circadian clock mutants of Synechococcus sp. Strain PCC 7942, selected transformed clones with kanamycin, screened cured clones which showed wildtype phenotype, and recovered the transferred genomic DNA segments from cured clones into E.ccli cells as plasmids by the plasmid rescue procedure. We identified a gene cluster kaiABC which is composed of three genes, kaiA, kaiB, and kaiC and is essential to the clock function (Ishiura et al., 1998. Science 281 : 1519-1523). All the mutations analyzed were missense mutations and mapped to this gene cluster. Homologs of kaiB and kaiC were found in the genomes of archaea, but not found in eukaryotes. Two promoters(PkaiA and PkaiBC), each one in both the upstream regions of kaiA and kaiB, were found, and kaiA and a set of kaiB and kaiC were transcribed as a kaiA mRNA and a kaiBC mRNA, respectively. The expression of both kaiA and kaiBC was circadian and affected by mutations in kaiA, kaiB, and kaiC themselves. KaiA protein enhanced the activity of PkaiBC promoter whereas KaiC protein repressed the activity. Therefore, the essence of the circadian clock in cyanobacteria is the circadian autoregulation of the expression of kaiABC by Kai proteins, KaiA, KaiB, and KaiC.

我们将基因组DNA文库转移到了Synechoccus sp的昼夜节律突变体中。菌株PCC 7942，选择了用卡纳霉素转化的克隆，筛选了固化的克隆，该克隆显示出野生型表型，并通过质粒救援程序从固定的克隆中恢复了从固化的克隆中转移的基因组DNA片段作为质粒，作为质粒。我们确定了一个基因簇kaiaBC，该基因由三个基因组成，kaia，kaib和kaic对时钟功能至关重要（Ishiura等，1998。Science281：1519-1523）。分析的所有突变都是错义突变，并映射到该基因簇。在古细菌的基因组中发现了KAIB和KAIC的同源物，但在真核生物中未发现。发现了两个启动子（PKAIA和PKAIBC），分别在Kaia和Kaib的上游区域中每个启动子，分别在Kaia和Kaib的上游区域中，Kaia和一组Kaib和Kaic分别作为Kaia mRNA和KAIBC mRNA转录。 Kaia和Kaibc的表达均为昼夜节律，并受到Kaia，Kaib和Kaic本身的突变的影响。 KAIA蛋白增强了PKAIBC启动子的活性，而KAIC蛋白抑制了活性。因此，蓝细菌中昼夜节律的本质是Kai蛋白，Kaia，Kaib和Kaic对Kaiabc表达的昼夜节律自动调节。

项目成果

Ishiura, M., Kutsuna, S., Aoki, S., Iwasaki, H., Andersson, C.R., Tanabe, A., Golden, S., Johnson, C.H., and Kondo, T.: "Expression of a gene cluster kaiABC as a circadianfeedback process in cyanobacteria." Science. 281. 1519-1523 (1998)

Ishiura, M.、Kutsuna, S.、Aoki, S.、Iwasaki, H.、Andersson, C.R.、Tanabe, A.、Golden, S.、Johnson, C.H. 和 Kondo, T.：“基因簇的表达

Tsinoremas,N.F.他: "A pigma factor that modifies the circadian expression of a subred of genes in cyanobactefia" EMBO.J.15. 2488-2495 (1996)

Tsinoremas, N.F. 等人：“一种改变蓝细菌亚红色基因昼夜节律表达的因子”EMBO.J.15 (1996)。

Tsinoremas et al.: "A sigma factor that rlifies the cricadian expression of a subeet of genes in yandoaecteria" EMBO.J.15. 2488-2495 (1996)

Tsinoremas 等人：“一种 sigma 因子，可增强 yandoaecteria 基因亚群的昼夜节律表达”EMBO.J.15。

Ishiura,M.,他: "Expression of a gene cluster kaiABC as a circadian feedback process in cyansbauterig." Science. 281. 1519-1523 (1998)

Ishiura, M., et al.：“作为昼夜节律反馈过程的基因簇 kaiABC 的表达，《科学》281. 1519-1523 (1998)。

Kondo, T., Mori, T., Lebedeva, N.V., Aoki, S., Ishiura, M., and Golden, S.S.: "Circadian rhythms in rapidly dividing cyanobacteria." Science. 275. 224-227 (1997)

Kondo, T.、Mori, T.、Lebedeva, N.V.、Aoki, S.、Ishiura, M. 和 Golden, S.S.：“快速分裂的蓝细菌的昼夜节律。”