A novel two‐component regulatory system in Bacillus subtilis for the survival of severe secretion stress

Abstract

The Gram-positive eubacterium Bacillus subtilis is well known for its high capacity to secrete proteins into the environment. Even though high-level secretion of proteins is an efficient process, it imposes stress on the cell. The present studies were aimed at the identification of systems required to combat this so-called secretion stress. A two-component regulatory system, named CssR-CssS, was identified, which bears resemblance to the CpxR-CpxA system of Escherichia coli. The results show that the CssR/S system is required for the cell to survive the severe secretion stress caused by a combination of high-level production of the alpha-amylase AmyQ and reduced levels of the extracytoplasmic folding factor PrsA. As shown with a prsA3 mutation, the Css system is required to degrade misfolded exported proteins at the membrane-cell wall interface. This view is supported by the observation that transcription of the htrA gene, encoding a predicted membrane-bound protease of B. subtilis, is strictly controlled by CssS. Notably, CssS represents the first identified sensor for extracytoplasmic protein misfolding in a Gram-positive eubacterium. In conclusion, the results show that quality control systems for extracytoplasmic protein folding are not exclusively present in the periplasm of Gram-negative eubacteria, but also in the Gram-positive cell envelope.

Membrane Proteases in the Bacterial Protein Secretion and Quality Control Pathway

“…In the enteric pathogen Vibrio cholerae, the causative agent of cholera, DegS is implicated in sigma E-mediated resistance to membrane-acting, host gut-derived antimicrobial peptides (103). In B. subtilis, the DegS homologue proteases HtrA and HtrB are both membrane bound by an N-terminal membrane anchor like E. coli DegS (7,19,62,198,199). Both enzymes have been implicated in the cellular response to the stresses caused by high-level production of secretory proteins or heat, but whether this involves a sheddase activity of HtrA or HtrB is not known (30,62,187).…”

“…In B. subtilis, the DegS homologue proteases HtrA and HtrB are both membrane bound by an N-terminal membrane anchor like E. coli DegS (7,19,62,198,199). Both enzymes have been implicated in the cellular response to the stresses caused by high-level production of secretory proteins or heat, but whether this involves a sheddase activity of HtrA or HtrB is not known (30,62,187). Interestingly, the cellular levels of HtrA and HtrB seem to be controlled by RasP, the RseP homologue of B. subtilis, suggesting that they are subject to intramembrane cleavage (19,199).…”

Heterologous production and secretion of Clostridium perfringens β-toxoid in closely related Gram-positive hosts

“…However, hardly any ␤-toxoid could be detected in the medium (data not shown). To test whether this could be due to the expression vector used, we tested the production and secretion of AmyQ, the alpha-amylase of B. amyloliquefaciens, which is widely used for testing secretion efficiencies in B. subtilis Hyyrylainen et al, 2001;Westers et al, 2004). The introduction of the vector with the rpsF promoter in front of amyQ (pRAT) resulted in high-level secretion of alpha-amylase.…”

The Bacillus subtilis NatK–NatR two-component system regulates expression of the natAB operon encoding an ABC transporter for sodium ion extrusion

“…Following the completion of genomic sequencing, several newly identified operons that encode two-component systems have been characterized, e.g. the BceRS, CitST, CssSR, DesKR, GlnKL, LiaRS, MalKR, YdbFG, YdfHI, YxdJK and YycFG systems (Aguilar et al, 2001;Asai et al, 2000;Doan et al, 2003;Fukuchi et al, 2000;Hyyrylainen et al, 2001;Joseph et al, 2004;Mascher et al, 2004;Ohki et al, 2003;Satomura et al, 2005;Serizawa & Sekiguchi, 2005;Tanaka et al, 2003;Yamamoto et al, 2000). In addition, close relationships have been identified between some operons encoding two-component systems and genes for ATP-binding cassette (ABC) transporter homologues, e.g.…”