title: The metalloprotease of Listeria monocytogenes controls cell wall translocation of the broad-range phospholipase C.
abstract: Listeria monocytogenes is a gram-positive bacterial pathogen that multiplies in the cytosol of host cells and spreads directly from cell to cell. During cell-to-cell spread, bacteria become temporarily confined to secondary vacuoles. The broad-range phospholipase C (PC-PLC) of L. monocytogenes contributes to bacterial escape from secondary vacuoles. PC-PLC requires cleavage of an N-terminal propeptide for activation, and Mpl, a metalloprotease of Listeria, is involved in the proteolytic activation of PC-PLC. Previously, we showed that cell wall translocation of PC-PLC is inefficient, resulting in accumulation of PC-PLC at the membrane-cell wall interface. In infected cells, rapid cell wall translocation of PC-PLC is triggered by a decrease in pH and correlates with cleavage of the propeptide in an Mpl-dependent manner. To address the role of the propeptide and of Mpl in cell wall translocation of PC-PLC, we generated a cleavage site mutant and a propeptide deletion mutant. The intracellular behavior of these mutants was assessed in pulse-chase experiments. We observed efficient translocation of the proform of the PC-PLC cleavage site mutant in a manner that was pH sensitive and Mpl dependent. However, the propeptide deletion mutant was efficiently translocated into host cells independent of Mpl and pH. Overall, these results suggest that Mpl regulates PC-PLC translocation across the bacterial cell wall in a manner that is dependent on the presence of the propeptide but independent of propeptide cleavage. In addition, similarly to Mpl-mediated cleavage of PC-PLC propeptide, Mpl-mediated translocation of PC-PLC across the bacterial cell wall is pH sensitive.
authors.affiliation: Department of Microbiology and Immunology, Cornell University, Ithaca, NY 14853-4601, USA.
authors.forename: P S Marie
authors.initials: PS
authors.lastname: Yeung
authors.forename: Nicholas
authors.initials: N
authors.lastname: Zagorski
authors.forename: Hélène
authors.initials: H
authors.lastname: Marquis
mesh_terms: D000818:Animals; D002460:Cell Line; D002473:Cell Wall / Q000378:metabolism*; D015964:Gene Expression Regulation, Bacterial; D006863:Hydrogen-Ion Concentration; D008089:Listeria monocytogenes / Q000201:enzymology* / Q000254:growth & development; D008264:Macrophages / Q000382:microbiology; D045726:Metalloproteases / Q000378:metabolism*; D051379:Mice; D010738:Type C Phospholipases / Q000378:metabolism*
chemical_list: D010738:Type C Phospholipases; C105598:phosphatidylcholine-specific phospholipase C; D045726:Metalloproteases