【作者】 王莉；

【导师】 张承才； 赵斌；

【作者基本信息】 华中农业大学 ， 微生物学， 2004， 博士

【摘要】 Anabaena PCC7120是一种丝状蓝细菌，在培养基中缺失化合态氮源的条件下，丝状体上分化形成异形胞执行固氮的功能。本论文主要的目的是研究异形胞发育过程中信号传导基因的功能。 Anabaena PCC7120的全基因组序列可以从CyanoBase数据库获得。基于其全基因组数据分析，发现了211个与双组份信号传导系统有关的基因，其中，有131个组氨酸激酶基因，80个反应调控子基因。根据这些基因的结构特征将它们分别分为7个亚家族和6个亚家族。分析显示，Anabaena PCC7120中有一些蛋白同时含有双组份系统和丝／苏氨酸激酶或磷酸酶的模件。其中，有一个由13个基因组成的亚家族，这个家族的蛋白既含有丝／苏氨酸激酶结构域也含有组氨酸激酶结构域。另外，还有4个基因编码的蛋白同时含有反应调控子结构域和丝／苏氨酸磷酸酶结构域。通过基因组比较分析发现，211个基因中约有35％的基因在Synechocystis PCC6803中可以找到同源基因，但其中有一个亚家族的组氨酸激酶基因和两个亚家族的反应调控子基因在Synechocystis PCC 6803中不存在任何同源基因。 本研究中共将109个基因进行了失活，其中19个基因获得了单交换突变株但未获得双交换突变株，74个获得了双交换突变株。在缺氮的条件下对双交换突变株分别进行表型观察。结果显示，有一个突变株不能分化异形胞；三个突变株可以形成异形胞，但在缺氮的条件下仍不能维持生长；4个突变株可以分化异形胞，但在缺氮条件下生长缓慢。这些基因可能在异形胞的分化或功能中发挥作用。 本研究还针对407个基因(包括信号传导有关的基因及转录因子)制备了基因芯片。利用基因芯片研究氮源缺失条件下基因表的达变化。结果显示，在缺氮培养24小时的条件下，28个基因的表达发生变化。以上结果为综合性研究异形胞的发育的分子机制提供了重要的分子生物学及遗传学的基础，便于更加深入的了解异形胞发育过程中的调控网络。更多还原

【Abstract】 Anabaena PCC 7120 is a filamentous cyanobacterium capable of both photosynthesis and dinitrogen fixation under aerobic conditions. Under conditions of combined nitrogen depletion, 5-10% of the cells along each fillament are induced to become specialized ones, called heterocysts, that are capable of molecular nitrogen fixation. Heterocyst create a microoxic environment to protect nitrogenase from oxygen. Up to now , the mechanism of singnal transduction involved in heterocyst differentiation is still unclear. The main objective of this study was to analyse genes of singnal transduction involved in heterocyst differentiation.Genome sequences of Anabaena PCC 7120 are available at the CyanoBase databank. Based on the genomic data, 211 genes encoding two-component signaling elements have been retrieved. There are 131 genes encoding histine kinase and 80 genes encoding response regulator, that are classified into 7 and 6 subfamilies respectively. Many proteins in Anabaena PCC 7120 have incorporated both modules of two-component systems and catalytic domains of either Ser/Thr kinases or phosphatases. A subfamily of 13 genes encode proteins with both a Ser/Thr kinase domain anti a His kinase domain, and another four genes were also found whose products have both a response regulator domain and a Ser/Thr phosphatase domain. Of all the signaling proteins in Anabaena PCC 7120, about one third (35%) are conserved in the genome of the unicellular cyanobacterium strain Synechocystis PCC 6803. Interestingly, one subfamily of His kinases and two subfamilies of response regulators are present in Anabaena PCC 7120, but absent in Synechocystis PCC 6803.109 genes were inactivated by insertion of a cassete confering antibiotic resistance. 19 single and 74 double recombinants were obtained. These mutants were examined for phenotypes after nitrogen depletion .The result showed that one mutant was unable to differentiate heterocyst, three formed heterocysts but still could not maintain normal growth, four growed slowly with heterocyst differentiation. These genes may play roles in hetetocyst differentiation or function.The gene chip used in this study contained 407 genes. The genes expression profil responding to nitrogen deprivation was analyzed using this gene-chip.The results indicated that 28 of 407 genes had an altered pattern of expression 24 hours after nitrogen deprivation. These studies provide a genetic and molecular basis for the understranding of the signaling network underlying heterocyst development in caynobacteria.更多还原