ABSTRACT
Bacterial cell wall contains peptidoglycan (PG) to protect the cells from turgor and environmental stress. PG consists of polymeric glycans cross-linked with each other by short peptide chains and forms an elastic mesh-like sacculus around the cytoplasmic membrane. Bacteria encode a plethora of PG hydrolytic enzymes of diverse specificity playing crucial roles in growth, division, or turnover of PG. In Escherichia coli, the cross-link-specific endopeptidases, MepS, -M, and -H, facilitate the enlargement of PG sacculus during cell elongation, whereas LytM-domain factors, EnvC and NlpD activate the division-specific amidases, AmiA, -B, and -C to facilitate the cell separation. In a screen to isolate additional factors involved in PG enlargement, we identified actS (encoding a LytM paralog, formerly ygeR) as its overexpression compensated the loss of elongation-specific endopeptidase, MepS. The overexpression of ActS resulted in the generation of partly denuded glycan strands in PG sacculi, indicating that ActS is either an amidase or an activator of amidase(s). The detailed genetic and biochemical analyses established that ActS is not a PG hydrolase, but an activator of the division-specific amidase, AmiC. However, interestingly, the suppression of the mepS growth defects by actS is not mediated through AmiC. The domain-deletion experiments confirmed the requirement of the N-terminal LysM domain of ActS for the activation of AmiC, but not for the alleviation of growth defects in mepS mutants, indicating that ActS performs two distinctive PG metabolic functions. Altogether our results suggest that in addition to activating the division-specific amidase, AmiC, ActS modulates yet another pathway that remains to be identified.
ABSTRACT
The bacterial cytoplasmic membrane is a principal site of protein translocation, lipid and peptidoglycan biogenesis, signal transduction, transporters and energy generating components of the respiratory chain. Although 25-30% of bacterial proteomes consist of membrane proteins, a comprehensive understanding of their influence on fundamental cellular processes is incomplete. Here, we show that YciB and DcrB, two small cytoplasmic membrane proteins of previously unknown functions, play an essential synergistic role in maintaining cell envelope integrity of Escherichia coli. Lack of both YciB and DcrB results in pleiotropic cell defects including increased levels of lipopolysaccharide, membrane vesiculation, dynamic shrinking and extension of the cytoplasmic membrane accompanied by lysis and cell death. The stalling of an abundant outer membrane lipoprotein, Lpp, at the periplasmic face of the inner membrane leads to lethal inner membrane-peptidoglycan linkages. Additionally, the periplasmic chaperone Skp contributes to yciB dcrB mutant cell death by possibly mistargeting stalled porins into the inner membrane. Consistent with the idea of a compromised envelope in the yciB dcrB mutant, multiple envelope stress response systems are induced, with Cpx signal transduction being required for growth. Taken together, our results suggest a fundamental role for YciB and DcrB in cell envelope biogenesis.
