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Essential PcsB putative peptidoglycan hydrolase interacts with the essential FtsXSpn cell division protein in Streptococcus pneumoniae D39.

ABSTRACT: The connection between peptidoglycan remodeling and cell division is poorly understood in ellipsoid-shaped ovococcus bacteria, such as the human respiratory pathogen Streptococcus pneumoniae. In S. pneumoniae, peptidoglycan homeostasis and stress are regulated by the WalRK (VicRK) two-component regulatory system, which positively regulates expression of the essential PcsB cysteine- and histidine-dependent aminohydrolases/peptidases (CHAP)-domain protein. CHAP-domain proteins usually act as peptidoglycan hydrolases, but purified PcsB lacks detectable enzymatic activity. To explore the functions of PcsB, its subcellular localization was determined. Fractionation experiments showed that cell-bound PcsB was located through hydrophobic interactions on the external membrane surface of pneumococcal cells. Immunofluorescent microscopy localized PcsB mainly to the septa and equators of dividing cells. Chemical cross-linking combined with immunoprecipitation showed that PcsB interacts with the cell division complex formed by membrane-bound FtsX(Spn) and cytoplasmic FtsE(Spn) ATPase, which structurally resemble an ABC transporter. Far Western blotting showed that this interaction was likely through the large extracellular loop of FtsX(Spn) and the amino terminal coiled-coil domain of PcsB. Unlike in Bacillus subtilis and Escherichia coli, we show that FtsX(Spn) and FtsE(Spn) are essential in S. pneumoniae. Consistent with an interaction between PcsB and FtsX(Spn), cells depleted of PcsB or FtsX(Spn) had strikingly similar defects in cell division, and depletion of FtsX(Spn) caused mislocalization of PcsB but not the FtsZ(Spn) early-division protein. A model is presented in which the interaction of the FtsEX(Spn) complex with PcsB activates its peptidoglycan hydrolysis activity and couples peptidoglycan remodeling to pneumococcal cell division.

SUBMITTER: Sham LT

PROVIDER: S-EPMC3215045 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature

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Essential PcsB putative peptidoglycan hydrolase interacts with the essential FtsXSpn cell division protein in Streptococcus pneumoniae D39.

Sham Lok-To LT Barendt Skye M SM Kopecky Kimberly E KE Winkler Malcolm E ME

Proceedings of the National Academy of Sciences of the United States of America 20111017 45

The connection between peptidoglycan remodeling and cell division is poorly understood in ellipsoid-shaped ovococcus bacteria, such as the human respiratory pathogen Streptococcus pneumoniae. In S. pneumoniae, peptidoglycan homeostasis and stress are regulated by the WalRK (VicRK) two-component regulatory system, which positively regulates expression of the essential PcsB cysteine- and histidine-dependent aminohydrolases/peptidases (CHAP)-domain protein. CHAP-domain proteins usually act as pepti ...[more]

PMID: 22006325

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Project description:Streptococcus pneumoniae is a human commensal bacterium that causes serious diseases. Peptidoglycan (PG) is the critical component of bacteria that maintains cell shape, size, chaining, and turgor resistance. Because of its surface exposure and eubacterial-specific mechanism of biosynthesis, PG biosynthesis remains an outstanding target for discovery of new antibiotics to resistant “superbugs,” like S. pneumoniae. Pneumococcal PG biosynthesis is carried out by a balance of septal and peripheral (side-wall-like) PG synthesis mediated by bPBP2x and bPBP2b, respectively, that emanates from midcell regions. We selected for additional suppressor mutations, besides mltG, that eliminate the requirement for essential bPBP2b in peripheral PG synthesis. We found that mutations that inactivate a putative RNA binding protein, designated KhpA, suppressed the requirement for some, but not all of the essential proteins required for peripheral PG synthesis. KhpA was used as bait in co-IP experiments to demonstrate interactions with a second RNA binding protein, designated KhpB, in cells. Mutations in khpA or khpB phenocopy each other and result in slower growth rates, smaller cell size with normal aspect ratios, and induction of cell wall stress responses. RIP-Seq analyses confirmed KhpAB as RNA binding protein in cells. Another suppressor of Δpbp2b implicated induction of a cell division operon as a way to bypass the requirement for bPBP2b. We show that increased expression of this cell division operon occurs in ΔkhpAB mutants and that this induction is necessary and sufficient to account for suppression of Δpbp2b. Comparisons of relative transcript amounts and protein levels indicate that induction of this cell division operon in ΔkhpAB mutants occurs primarily at the post-transcriptional level. Together, our results show that KhpAB, which are virulence factors in S. pneumoniae and are conserved in other Gram-positive pathogens, acts as a new class of RNA binding protein, with properties analogous to Hfq in Gram-negative bacteria.

Dataset Information

Essential PcsB putative peptidoglycan hydrolase interacts with the essential FtsXSpn cell division protein in Streptococcus pneumoniae D39.

Publications

Essential PcsB putative peptidoglycan hydrolase interacts with the essential FtsXSpn cell division protein in Streptococcus pneumoniae D39.

Similar Datasets

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