Transcriptional Program of Early Sporulation and Stationary-Phase Events in Clostridium acetobutylicum

Abstract

DNA microarray analysis of Clostridium acetobutylicum was used to examine the genomic-scale gene expression changes during the shift from exponential-phase growth and acidogenesis to stationary phase and solventogenesis. Self-organizing maps were used to identify novel expression patterns of functional gene classes, including aromatic and branched-chain amino acid synthesis, ribosomal proteins, cobalt and iron transporters, cobalamin biosynthesis, and lipid biosynthesis. The majority of pSOL1 megaplasmid genes (in addition to the solventogenic genes aad-ctfA-ctfB and adc) had increased expression at the onset of solventogenesis, suggesting that other megaplasmid genes may play a role in stationary-phase phenomena. Analysis of sporulation genes and comparison with published Bacillus subtilis results indicated conserved expression patterns of early sporulation genes, including spo0A, the sigF operon, and putative canonical genes of the H and F regulons. However, sigE expression could not be detected within 7.5 h of initial spo0A expression, consistent with the observed extended time between the appearance of clostridial forms and endospore formation. The results were compared with microarray comparisons of the wild-type strain and the nonsolventogenic, asporogenous M5 strain, which lacks the pSOL1 megaplasmid. While some results were similar, the expression of primary metabolism genes and heat shock proteins was higher in M5, suggesting a difference in metabolic regulation or a butyrate stress response in M5. The results of this microarray platform and analysis were further validated by comparing gene expression patterns to previously published Northern analyses, reporter assays, and two-dimensional protein electrophoresis data of metabolic genes (including all major solventogenesis genes), sporulation genes, heat shock proteins, and other solventogenesis-induced gene expression.Newly developed genomic technologies allow high-throughput screening of entire transcriptomes. Several recent studies of Bacillus subtilis have utilized the DNA microarray technology for the analysis of key mutants to identify genes affected by sporulation in addition to identifying new genes in sporulation regulons (8,18,19,22). A fundamental biological problem is to identify gene expression patterns for all major functional classes during the shift from exponential-phase growth to stationary phase, especially in endospore-forming organisms such as the bacilli and clostridia, and relate such changes to important physiological changes and events. The number of affected genes is potentially very large; in B. subtilis, it has been shown that inactivation of the stationary-phase gene regulator spo0A can directly and indirectly affect the expression of approximately 500 genes at least threefold during the onset of sporulation (22).Solventogenic clostridia like Clostridium acetobutylicum produce acids during exponential-phase growth, and during stationary phase, they form granulose, take up acids to produce solvents, and sporulate. An understan...

Degradation of ppGpp by Nudix Pyrophosphatase Modulates the Transition of Growth Phase in the Bacterium Thermus thermophilus

“…suppression of the tricarboxylic acid cycle, cobalamin biosynthesis, DNA replication, and induction of stress response proteins (43)(44)(45)(46). The transcriptional suppression of the septum formation component FtsZ and DNA replication machinery DnaB could be directly attributed to the cellular elongation and growth inhibition for the mutant (47,48).…”

The putative influence of the agr operon upon survival mechanisms used by Clostridium acetobutylicum

“…In S. aureus, this leads to upregulation of all four agr genes. In C. acetobutylicum, on the other hand, although adjacent, agrBD and agrCA are predicted to form independent transcriptional units [31] and, accordingly, transcriptome data of [17] illustrate that only transcripts of agrBD are significantly increased at stationary phase. Thus the evidence suggests that quorum sensing feedback into agrCA is absent in C. acetobutylicum.…”

“…Previous modelling studies have suggested that feedback into the two-component system is the key to a fully effective quorum-sensing system [16], but, interestingly, these two feedback loops appear to be absent in C. acetobutylicum [17]. Comparisons with the staphylococcal agr system indicate that, depending upon the mechanism of interaction between the agr system and the sporulation-initiation network, the clostridial agr circuitry may (at least in part) be in place either to moderate the number of spores that are formed (in order for this number to reflect the urgency of the situation), or simply as an energy-saving strategy.…”

Engineering Complex Phenotypes in Industrial Strains

“…Association analysis of both sets of data, transcriptional and metabolite profiles, led to the identification of targets for further metabolic engineering. Microarray-based transcriptome analysis in engineering solvent (butanol) tolerance (71,72) and identification of genetic determinants contributing to a variety of phenotypes is reported in the literature (68,(73)(74)(75)(76)(77)(78)(79)(80)(81)(82).…”