CIRCADIAN-RHYTHMS IN PROKARYOTES - LUCIFERASE AS A REPORTER OF CIRCADIAN GENE-EXPRESSION IN CYANOBACTERIA
CIRCADIAN-RHYTHMS IN PROKARYOTES - LUCIFERASE AS A REPORTER OF CIRCADIAN GENE-EXPRESSION IN CYANOBACTERIA
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DOI：
10.1073/pnas.90.12.5672
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发表时间：
1993-06-15
影响因子：
11.1
通讯作者：
JOHNSON, CH
中科院分区：
文献类型：
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作者：
KONDO, T;STRAYER, CA;JOHNSON, CH
我们已经使用了荧光素酶报告基因和生物发光的连续自动化监测，以明确地表明，蓝藻表现出的昼夜节律的行为，从根本上说是相同的真核生物的昼夜节律。我们还表明，这些节奏可以通过分子生物学方法研究聚球藻。PCC 7942，一个菌株，其中遗传转化是建立。将哈氏弧菌荧光素酶结构基因（luxAB）的无启动子区段引入聚球藻psbAI基因的启动子下游，所述聚球藻psbAI基因编码光系统II蛋白。将该报告构建重组到聚球藻染色体中，并在夹带到光和暗循环后的恒定照明条件下监测生物发光。报告菌株，AMC 149，表达了生物发光的节奏，满足昼夜节律的标准：在恒定条件下的持久性，通过光/暗信号的相位重置，和温度补偿期间。光/暗夹带后天然psbAI信息水平的节律性变化支持报告者数据。这种原核生物的行为反驳了昼夜节律机制必须基于真核细胞组织的教条。此外，这里描述的蓝藻菌株提供了一个有效的实验系统的生物钟的分子分析。
We have used a luciferase reporter gene and continuous automated monitoring of bioluminescence to demonstrate unequivocally that cyanobacteria exhibit circadian behaviors that are fundamentally the same as circadian rhythms of eukaryotes. We also show that these rhythms can be studied by molecular methods in Synechococcus sp. PCC7942, a strain for which genetic transformation is well established. A promoterless segment of the Vibrio harveyi luciferase structural genes (luxAB) was introduced downstream of the promoter for the Synechococcus psbAI gene, which encodes a photosystem II protein. This reporter construction was recombined into the Synechococcus chromosome, and bioluminescence was monitored under conditions of constant illumination following entrainment to light and dark cycles. The reporter strain, AMC149, expressed a rhythm of bioluminescence which satisfies the criteria of circadian rhythms: persistence in constant conditions, phase resetting by fight/dark signals, and temperature compensation of the period. Rhythmic changes in levels of the native psbAI message following light/dark entrainment supported the reporter data. The behavior of this prokaryote disproves the dogma that circadian mechanisms must be based on eukaryotic cellular organization. Moreover, the cyanobacterial strain described here provides an efficient experimental system for molecular analysis of the circadian clock.