The dynamic architecture of the metabolic switch in Streptomyces coelicolor

Abstract

Background: During the lifetime of a fermenter culture, the soil bacterium S. coelicolor undergoes a major metabolic switch from exponential growth to antibiotic production. We have studied gene expression patterns during this switch, using a specifically designed Affymetrix genechip and a high-resolution time-series of fermenter-grown samples.

Results: Surprisingly, we find that the metabolic switch actually consists of multiple finely orchestrated switching events. Strongly coherent clusters of genes show drastic changes in gene expression already many hours before the classically defined transition phase where the switch from primary to secondary metabolism was expected. The main switch in gene expression takes only 2 hours, and changes in antibiotic biosynthesis genes are delayed relative to the metabolic rearrangements. Furthermore, global variation in morphogenesis genes indicates an involvement of cell differentiation pathways in the decision phase leading up to the commitment to antibiotic biosynthesis.

Conclusions: Our study provides the first detailed insights into the complex sequence of early regulatory events during and preceding the major metabolic switch in S. coelicolor, which will form the starting point for future attempts at engineering antibiotic production in a biotechnological setting.

Figure 1

Characterization of the fermentation time series samples. A. Major biochemical parameters of the fermentation run. The figure shows the increase in biomass (brown), phosphate depletion (pink), the decrease of glucose (yellow) and glutamate (green) levels, as well as the production of the antibiotics undecylprodigiosin (red), actinorhodin (light blue), and total blue pigment (dark blue). Continuous measurements of CO₂ production are indicated by the thin red line. The sampling intervals for the gene expression measurements are indicated at the top. B. Hierarchical clustering of the 32 samples along the time course. Each leaf of the tree corresponds to one sample and the labels indicate the time (in hours) after inoculation at which the sample was harvested. The clustering of the 32 samples is based on the expression profiles of 322 transcripts with highest regularized variance. A neighbor joining tree was produced from the pairwise Euclidean distances between the expression profiles of all samples.

Figure 2

Representative genes of the ribosomal gene cluster, showing continuous decrease in expression. The major expression change happens after 35 h, when phosphate is depleted in the medium (grey vertical line). A list of the genes and their expression data is included in Additional File 2.

Figure 3

Representative genes of the nitrogen metabolism gene cluster. A clear switch is visible already at 24 h, and after 31 h the genes are approaching background expression levels. A list of the genes and their expression data is included in Additional File 2.

Figure 4

Representative genes of the CPK antibiotics gene cluster. A strong transient expression peak is seen around 24 h, for all of the genes, and many of them remain at constantly elevated expression levels afterwards. A list of the genes and their expression data is included in Additional File 2.

Figure 5

Representative genes of the cluster of development related genes. These genes, which are involved in morphogenesis and the sporulation process, show a continuous increase in expression early in the time course, but are repressed immediately before the major metabolic switch happens. A list of the genes and their expression data is included in Additional File 2.

Figure 6

Genes upregulated by phosphate depletion. These include the regulatory genes phoP, phoU and phoR, genes of the phosphate transport cluster (pstABCS), as well as 13 other genes identified as activated by PhoP in the pho regulon [20-22]. The dramatic change in expression of these genes coincides exactly with the major metabolic switch induced by phosphate depletion. A list of the genes and their expression data is included in Additional File 2.

Figure 7

Representative genes involved in synthesis of phosphate-free secondary polymers of the cell-wall that closely follow the expression profile of the phosphate transporters. These genes are all part of a single genomic cluster (SCO4873-4882) and are part of the pho regulon. A list of the genes and their expression data is included in Additional File 2.

Figure 8

Expression profile of the Red and Act antibiotics biosynthesis gene cluster. The expression change of these genes, as a consequence of the metabolic switch, leads to the production of pigment antibiotics that characterizes the stationary growth phase of Streptomyces. Their upregulation is shifted by many hours, relative to the phosphate starvation, indicating a systematic delay in their regulatory mechanism. The gene expression levels are closely correlated to the increase in antibiotics levels measured in the medium. A list of the genes and their expression data is included in the supplementary table. Upper plot: Red cluster; lower plot: Act cluster.

Figure 9

Sequence of expression switches associated with the metabolic switch. The expression profile of the ten most variable transcriptional regulators on the array is shown. As discussed in the main text, some of these regulators show an expression profile that clearly parallels that of the major expression clusters. These regulators are the main candidate drivers of the successive waves of gene expression that characterize the metabolic switch according to our study. They include phosphate transport system regulators (phoP, dark grey, phoU, light green, and phoR, dark green), the transcriptional regulator of the red cluster, redD (red), the pyrimidine regulatory protein pyrR (blue), as well as the nitrogen regulators glnR (light purple) and glnK (dark purple) [10]. A list of the genes and their expression data is included in Additional File 2.