Global transcriptomic analysis of Cyanothece 51142 reveals robust diurnal oscillation of central metabolic processes

Abstract

Cyanobacteria are photosynthetic organisms and are the only prokaryotes known to have a circadian lifestyle. Unicellular diazotrophic cyanobacteria such as Cyanothece sp. ATCC 51142 produce oxygen and can also fix atmospheric nitrogen, a process exquisitely sensitive to oxygen. To accommodate such antagonistic processes, the intracellular environment of Cyanothece oscillates between aerobic and anaerobic conditions during a day-night cycle. This is accomplished by temporal separation of the two processes: photosynthesis during the day and nitrogen fixation at night. Although previous studies have examined periodic changes in transcript levels for a limited number of genes in Cyanothece and other unicellular diazotrophic cyanobacteria, a comprehensive study of transcriptional activity in a nitrogen-fixing cyanobacterium is necessary to understand the impact of the temporal separation of photosynthesis and nitrogen fixation on global gene regulation and cellular metabolism. We have examined the expression patterns of nearly 5,000 genes in Cyanothece 51142 during two consecutive diurnal periods. Our analysis showed that approximately 30% of these genes exhibited robust oscillating expression profiles. Interestingly, this set included genes for almost all central metabolic processes in Cyanothece 51142. A transcriptional network of all genes with significantly oscillating transcript levels revealed that the majority of genes encoding enzymes in numerous individual biochemical pathways, such as glycolysis, oxidative pentose phosphate pathway, and glycogen metabolism, were coregulated and maximally expressed at distinct phases during the diurnal cycle. These studies provide a comprehensive picture of how a physiologically relevant diurnal light-dark cycle influences the metabolism in a photosynthetic bacterium.

Fig. 1.

Cycling of gene expression in Cyanothece 51142. (A) The percentage of cycling genes that peak at dark–light and light–dark transitions is nearly twice as high as at other times. Percentages are of all cycling genes. (B) Relative cellular transcript abundance over time shows a maximum at D5, corresponding with peaks in nitrogenase-related gene expression (see Fig. 2A). The alternating dark–light cycles are indicated as black and white bars below the x axis.

Fig. 2.

Expression profiles of genes involved in (A) nitrogen fixation and (B) genes encoding ribosomal proteins. Dark–light cycles are indicated as black and white bars below the x-axis. Time points are designated as hours exposed to dark or light. The log₂ ratios of transcript abundance to the pooled sample control are plotted on the y axis.

Fig. 3.

Coexpression network of strongly cycling genes. Genes whose transcript levels changed by at least 2-fold over the entire time course were visualized by using Cytoscape version 2.3.2 (21). Genes with Pearson correlation coefficients ≥0.9 are connected, and each node corresponds to one gene. Genes are colored according to their transcript abundance at each time point: red and yellow, genes up-regulated by 4- and 2-fold, respectively; green, genes not differentially expressed; cyan, blue, and light blue, genes down-regulated by 2-, 4-, and 16-fold, respectively. The first 24 h of the 48-h time course are shown. The four different subnetworks are labeled according to the most prominent functional category.

Fig. 4.

Schematic organization of a Cyanothece cell showing central metabolic pathways. Pathways are divided according to the diurnal period in which they are most up-regulated. Double arrows show pathways that involve reversible reactions. The percentage of up-regulated enzymes in each pathway is in red, with the percentage of all up-regulated isoforms in parentheses.