Glucose-Dependent Activation ofBacillus anthracisToxin Gene Expression and Virulence Requires the Carbon Catabolite Protein CcpA

Abstract

Sensing environmental conditions is an essential aspect of bacterial physiology and virulence. In Bacillus anthracis, the causative agent of anthrax, transcription of the two major virulence factors, toxin and capsule, is triggered by bicarbonate, a major compound in the mammalian body. Here it is shown that glucose is an additional signaling molecule recognized by B. anthracis for toxin synthesis. The presence of glucose increased the expression of the protective antigen toxin component-encoding gene (pagA) by stimulating induction of transcription of the AtxA virulence transcription factor. Induction of atxA transcription by glucose required the carbon catabolite protein CcpA via an indirect mechanism. CcpA did not bind specifically to any region of the extended atxA promoter. The virulence of a B. anthracis strain from which the ccpA gene was deleted was significantly attenuated in a mouse model of infection. The data demonstrated that glucose is an important host environment-derived signaling molecule and that CcpA is a molecular link between environmental sensing and B. anthracis pathogenesis.The capacity for sensing and responding to their surroundings is an essential feature of all living organisms. Bacteria have developed an array of highly sophisticated sensing and adaptation systems, such as two-component systems, quorum-sensing systems, and transport systems (9,15,21,42,58). Investigations into bacterial pathogenesis have revealed an intuitive connection between basic metabolic processes of adaptation and virulence. For example, it has been reported that bacteria alter the transcription of carbohydrate utilization genes and virulence factor-encoding genes in response to the sugar availability encountered in the environment, in particular, in the infected host (44,65,67,69,79). Consequently, regulatory mechanisms that connect carbohydrate metabolism to virulence factor production must have evolved.A key role in carbohydrate utilization in both Gram-negative and Gram-positive organisms is played by the complex sugartransporting phosphotransferase system (PTS), but additional proteins and different mechanisms are involved in the two types of bacteria (4,16,53). One of these mechanisms is the carbon catabolite repression (CCR) system that is based on the Crp-cyclic AMP system in Gram-negative bacteria, while the HPr protein of the PTS system, together with the transcription factor CcpA, is the master regulator in Gram-positive bacteria (27,59,75).Extensive work carried out mostly in Bacillus subtilis has demonstrated that HPr can be phosphorylated on the His15 residue (B. subtilis numeration) by the enzyme I (EI) of the PTS in response to the phosphoenolpyruvate-to-pyruvate ratio or on the Ser46 residue by the HPr kinase/phosphorylase (HPrK/P) enzyme in response to the concentrations of ATP, P i , PP i , and fructose-1,6-bisphosphate (FBP) (23,30,43,56). HPr phosphorylated on Ser46 interacts and stimulates CcpA DNA binding activity, thus providing the link between carbon availability and transcriptional a...

cis -Acting Elements That Control Expression of the Master Virulence Regulatory Gene atxA in Bacillus anthracis

“…In agreement with the significance of AtxA in pathogenesis, atxA transcript levels are 5-to 6-fold greater in cultures grown at 37°C than in cultures incubated at 30°C (10). In the presence of glucose, the catabolite control protein CcpA stimulates transcription of atxA indirectly by an unknown mechanism (9). The redox state of the cell also appears to control atxA transcription.…”

A Novel Mode of Regulation of the Staphylococcus aureus Catabolite Control Protein A (CcpA) Mediated by Stk1 Protein Phosphorylation

“…The CcpA⅐Hpr-Ser(P)-46 complex has an increased affinity for particular cis-acting sequences, termed catabolite-responsive element (cre) sequences, and thereby represses or enhances gene expression depending on the position of the cre in relation to the operator sequence (3,4). In a number of Gram-positive pathogens such as Bacillus anthracis (5), Clostridium difficile (6), Staphylococcus aureus (7), Staphylococcus epidermidis (8), Streptococcus pneumoniae (9), and Streptococcus pyogenes (10), CcpA is also involved in the regulation of virulence determinants.…”

“…The corresponding amplified product was then digested with NheI and BamHI and ligated into the pETPhos plasmid, thus generating pETPhos_ccpA Ba , which was transformed and overexpressed, and the His-tagged CcpA was purified as described above. The B. anthracis ccpA recombinant vector was a kind gift from Marta Perego and purified as described by Chiang et al (5) to generate CcpA Ba .…”

Simple detection of Bacillus anthracis spores by precipitation method with goat antibody anti anthrosa

“…It has been shown that induction of Bacillus anthracis toxin gene expression by glucose and determined that CcpA transcription factor plays a positive role by indirectly regulating the transcription of the gene encoding AtxA. 31 Bacillus anthracis cultured on agar with 7% NaCl showed good growth (Figure 1 (F1)). We suspect, the nature of which is caused by the structure of the cell membrane that are typical of Bacillus anthracis.…”

“…Further also been shown that glucose is an additional signaling molecule recognized by Bacillus anthracis for toxin synthesis. 31 Bacillus anthracis cultured in nutrient broth when shaken showed the typical image that is like tendrils of silkworms (Figure 1 (C1)), while Bacillus cereus ( Figure 1 (C2)) and Bacillus thuringiensis (Figure 1 (C3)) remain murky. The fact is the hallmark of Bacillus anthracis.…”