title: Decrease in penicillin susceptibility due to heat shock protein ClpL in Streptococcus pneumoniae.
abstract: Antibiotic resistance and tolerance are increasing threats to global health as antibiotic-resistant bacteria can cause severe morbidity and mortality and can increase treatment cost 10-fold. Although several genes contributing to antibiotic tolerance among pneumococci have been identified, we report here that ClpL, a major heat shock protein, could modulate cell wall biosynthetic enzymes and lead to decreased penicillin susceptibility. On capsular type 1, 2, and 19 genetic backgrounds, mutants lacking ClpL were more susceptible to penicillin and had thinner cell walls than the parental strains, whereas a ClpL-overexpressing strain showed a higher resistance to penicillin and a thicker cell wall. Although exposure of Streptococcus pneumoniae D39 to penicillin inhibited expression of the major cell wall synthesis gene pbp2x, heat shock induced a ClpL-dependent increase in the mRNA levels and protein synthesized by pbp2x. Inducible ClpL expression correlated with PBP2x expression and penicillin susceptibility. Fractionation and electron micrograph data revealed that ClpL induced by heat shock is localized at the cell wall, and the ΔclpL showed significantly reduced net translocation of PBP2x into the cell wall. Moreover, coimmunoprecipitation with either ClpL or PBP2x antibody followed by reprobing with ClpL or PBP2x antibody showed an interaction between ClpL and PBP2x after heat stress. This interaction was confirmed by His tag pulldown assay with either ClpLHis₆ or PBP2xHis₆. Thus, ClpL stabilized pbp2x expression, interacted with PBP2x, and facilitated translocation of PBP2x, a key protein of cell wall synthesis process, contributing to the decrease of antibiotic susceptibility in S. pneumoniae.
authors.affiliation: School of Pharmacy, Sungkyunkwan University, Suwon 440-746, South Korea.
authors.forename: Thao Dang-Hien
authors.initials: TD
authors.lastname: Tran
authors.forename: Hyog-Young
authors.initials: HY
authors.lastname: Kwon
authors.forename: Eun-Hye
authors.initials: EH
authors.lastname: Kim
authors.forename: Ki-Woo
authors.initials: KW
authors.lastname: Kim
authors.forename: David E
authors.initials: DE
authors.lastname: Briles
authors.forename: Suhkneung
authors.initials: S
authors.lastname: Pyo
authors.forename: Dong-Kwon
authors.initials: DK
authors.lastname: Rhee
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics; D001433:Bacteriolysis / Q000187:drug effects; D003840:Deoxycholic Acid / Q000494:pharmacology; D024881:Drug Resistance, Bacterial; D006360:Heat-Shock Proteins / Q000502:physiology*; D006358:Hot Temperature; D017830:Octoxynol / Q000494:pharmacology; D010403:Penicillin Resistance; D046915:Penicillin-Binding Proteins / Q000235:genetics / Q000378:metabolism; D010406:Penicillins / Q000494:pharmacology; D010453:Peptide Synthases / Q000235:genetics; D013296:Streptococcus pneumoniae / Q000187:drug effects* / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D006360:Heat-Shock Proteins; D046915:Penicillin-Binding Proteins; D010406:Penicillins; D003840:Deoxycholic Acid; C072223:PBP 2x protein, Streptococcus; D017830:Octoxynol; C408066:MurM protein, Streptococcus pneumoniae; D010453:Peptide Synthases