PMID:18599819

Citation	Wang, L and Lin, M (2008) A novel cell wall-anchored peptidoglycan hydrolase (autolysin), IspC, essential for Listeria monocytogenes virulence: genetic and proteomic analysis. Microbiology (Reading, Engl.) 154:1900-13
Abstract	We have recently concluded that a Listeria monocytogenes 86 kDa immunogenic surface protein, IspC, is a cell wall-anchored peptidoglycan hydrolase (autolysin), capable of degrading the cell wall peptidoglycan of the bacterium itself. To determine if this enzyme has any biological functions and/or plays a role in virulence, we in-frame-deleted the ispC gene from the L. monocytogenes chromosome. This DeltaispC mutant exhibited complete abrogation of expression of IspC and displayed no defects in in vitro growth, colony and microscopic morphologies, or biochemical characteristics. Lack of IspC led to attenuated virulence in mice, evidenced by a significant reduction in bacterial counts in livers and brains and no mortality compared with the wild-type. Furthermore, the data from assays using various eukaryotic cells for adhesion, invasion, actin tail formation, plaque formation and intracellular growth indicated that the mutant was severely attenuated in virulence in a cell culture model in a cell type-dependent manner. The findings that (i) the mutant was impaired for adhesion to certain eukaryotic cells, and (ii) both purified IspC and its C-terminal cell wall-binding domain were capable of binding sheep choroid plexus (SCP) epithelial cells and Vero cells, supported the role of IspC as an adhesin in virulence. The DeltaispC mutant exhibited a marked defect in adhesion to and invasion of SCP cells but not human brain microvascular endothelial cells (HBMEC), suggesting that IspC is necessary for crossing the blood-cerebrospinal fluid barrier. Proteomic and immunological analysis showed a reduced surface expression of some known or putative virulence factors (e.g. ActA, InlC2 and a flagellin homologue, FlaA) due to IspC deficiency. Altogether, this study demonstrates that IspC, expressed as a minor autolysin in vitro, is not important for cell division or separation but is essential for full virulence of L. monocytogenes in vivo.
Links	PubMed Online version:10.1099/mic.0.2007/015172-0
Keywords	Animals; Bacterial Adhesion; Bacterial Proteins/chemistry; Bacterial Proteins/genetics; Bacterial Proteins/metabolism; Cell Line, Tumor; Cell Wall/enzymology; Cell Wall/genetics; Cell Wall/physiology; Cercopithecus aethiops; Choroid Plexus/microbiology; Endothelial Cells/microbiology; Gene Expression Regulation, Bacterial; Humans; Listeria monocytogenes/enzymology; Listeria monocytogenes/genetics; Listeria monocytogenes/pathogenicity; Listeria monocytogenes/physiology; Listeriosis/microbiology; Mice; Mice, Inbred BALB C; N-Acetylmuramoyl-L-alanine Amidase/chemistry; N-Acetylmuramoyl-L-alanine Amidase/genetics; N-Acetylmuramoyl-L-alanine Amidase/metabolism; Phenotype; Protein Binding; Protein Structure, Tertiary; Proteomics; Sequence Deletion; Sheep; Species Specificity; Vero Cells; Virulence; Virulence Factors/genetics; Virulence Factors/metabolism
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Annotations

Gene product	Qualifier	GO Term	Evidence Code	with/from	Aspect	Extension	Notes	Status
LISMN:A3RI35	involved_in	GO:0044650: adhesion of symbiont to host cell	ECO:0000315: mutant phenotype evidence used in manual assertion		P		Seeded From UniProt	complete
LISMN:A3RI35		GO:0044650: adhesion of symbiont to host cell	ECO:0000315:		P		Fig 3 shows listeria monocytogenes cells carrying a deletion mutation of the ispC gene have decreased adherence to non-phagocytic eukaryotic cell lines as compared to the wild-type strain.	complete CACAO 9877

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