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Comprehensive study on transcriptional network system in the photoautotrophic cyanobacteria Synechococcusstrains

光合自养蓝细菌聚球藻转录网络系统的综合研究

基本信息

批准号：
13206027
负责人：
SUGITA Mamoru
金额：
$ 37.31万
依托单位：
Nagoya University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research on Priority Areas
财政年份：
2001
资助国家：
日本
起止时间：
2001 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-13206027/
关键词：
cyanobacteria transcription network two-component regulatory system nitrate transport system DNA chip photoautotroph circadian rhythm clock gene ゲノム光合成遺伝子炭酸水素イオン能動輸送体 Synechococcus 6301株 Synechococcus 7942株 Synechocystis 6803株

项目摘要

(1) The entire genome of a unicellular cyanobacterium, Synechococcus sp. strain PCC 6301, was sequenced. The genome consisted of a circular chromosome 2,696,255 bp long. A total of 2525 potential protein-encoding genes, two sets of rRNA genes, 45 tRNA genes representing 42 tRNA species and several genes, for small structural RNAs were assigned to the chromosome. Thirty-seven genes for the proteins involved in two-component regulatory system were also annotated. Ten percent of all the protein genes lacked significant similarity to genes for predicted proteins in the public, indicating the genes unique to Synechococcus PCC6301. We constructed high-density DNA microarrays (Affimetrix Gene Chip) of Synechococcus PCC6301 as a tool for comprehensive analysis of transcriptional network in cyanobacteria.(2) We identified three genes responsible for the latent transport activity for nitrate: ltnA, which encodes a response regulator with no effector domain; ltnB, which encodes a hybrid histidine kinase with two receiver domains; and ltnT, which encodes a sulfate permease-like protein with a putative cyclic nucleoside monophosphate (cNMP)-binding domain.(3) Circadian clock of cyanobacteria was considered to be composed of negative feedback regulation of kaiBC expression. We demonstrated that KaiC phosphorylation state oscillated even without kaiBC messenger RNA accumulation under continuous dark conditions. Moreover, kinetic profiles in the ratio of KaiC autophosphorylation-dephosphorylation were also temperature compensated in vitro. Thus, the cyanobacterial clock can keep time independent of de novo transcription and translation processes. Furthermore, we have reconstituted the self-sustainable oscillation of KaiC phosphorylation in vitro. The period of the in vitro oscillation was stable and the circadian periods observed in vivo in KaiC mutant strains were consistent with those measured in vitro.

(1)对单细胞蓝细菌聚球藻属菌株PCC 6301的全基因组进行了测序。基因组由一条长2，696，255 bp的环状染色体组成。共有2525个潜在的蛋白质编码基因，两套rRNA基因，45个tRNA基因代表42种tRNA和几个基因，小结构RNA被分配到染色体。对37个参与双组分调控系统的蛋白质基因进行了注释。10%的蛋白质基因与公众预测的蛋白质基因缺乏显著的相似性，表明聚球藻PCC 6301特有的基因。我们构建了聚球藻PCC 6301的高密度DNA微阵列（Affiliation Gene Chip），作为全面分析蓝藻转录网络的工具。(2)我们确定了三个基因负责硝酸盐的潜在运输活性：ltnA，它编码一个没有效应域的响应调节器; ltnB，它编码一个有两个接收器域的混合组氨酸激酶;和ltnT，它编码一个硫酸通透酶样蛋白，具有一个假定的环核苷一磷酸（cNMP）结合域。(3)蓝藻的生物钟被认为是由kaiBC表达的负反馈调节组成的。我们证明，即使在连续黑暗条件下没有kaiBC信使RNA积累，KaiC磷酸化状态也会振荡。此外，KaiC自磷酸化-去磷酸化比率的动力学曲线在体外也是温度补偿的。因此，蓝藻时钟可以保持时间独立的从头转录和翻译过程。此外，我们在体外重建了KaiC磷酸化的自我维持振荡。在体外振荡的周期是稳定的，在体内观察到的KaiC突变株的昼夜节律周期与在体外测量的一致。