Identifying the components of theShewanellaphage LambdaSo lysis system

Abstract

Phage-induced lysis of Gram-negative bacterial hosts usually requires a set of phage lysis proteins, a holin, an endopeptidase and a spanin system, to disrupt each of the three cell envelope layers. Genome annotations and previous studies identified a gene region in theShewanella oneidensisprophage LambdaSo, which comprises potential holin- and endolysin-encoding genes but lacks an obvious spanin system. By a combination of candidate approaches, mutant screening, characterization and microscopy we found that LambdaSo uses a pinholin/signal-anchor-release (SAR) endolysin system to induce proton-leakage and degradation of the cell wall. Between the corresponding genes we found that two extensively nested open reading frames encode a two-component spanin module Rz/Rz1. Unexpectedly, we identified another factor strictly required for LambdaSo-induced cell lysis, the phage protein Lcc6. Lcc6 is a transmembrane protein of 65 amino acid residues with hitherto unknown function, which acts at the level of holin in the cytoplasmic membrane to allow endolysin release. Thus, LambdaSo-mediated cell lysis requires at least four protein factors (pinholin, SAR-endolysin, spanin, Lcc6). The findings further extend the known repertoire of phage proteins involved in host lysis and phage egress.