Abstract
ABSTRACTThe chitinolytic system of Listeria monocytogenes thus far comprises two chitinases, ChiA and ChiB, and a lytic polysaccharide monooxygenase, Lmo2467. The role of the system in the bacterium appears to be pleiotropic, as besides mediating the hydrolysis of chitin, the second most ubiquitous carbohydrate in nature, the chitinases have been deemed important for the colonization of unicellular molds, as well as mammalian hosts. To identify additional components of the chitinolytic system, we screened a transposon mutant library for mutants exhibiting impaired chitin hydrolysis. The screening yielded a mutant with a transposon insertion in a locus corresponding to lmo0327 of the EGD-e strain. lmo0327 encodes a large (1,349 amino acids [aa]) cell wall-associated protein that has been proposed to possess murein hydrolase activity. The single inactivation of lmo0327, as well as of lmo0325 that codes for a putative transcriptional regulator functionally related to lmo0327, led to an almost complete abolishment of chitinolytic activity. The effect could be traced at the transcriptional level, as both chiA and chiB transcripts were dramatically decreased in the lmo0327 mutant. In accordance with that, we could barely detect ChiA and ChiB in the culture supernatants of the mutant strain. Our results provide new information regarding the function of the lmo0325-lmo0327 locus in L. monocytogenes and link it to the expression of chitinolytic activity.IMPORTANCE Many bacteria from terrestrial and marine environments express chitinase activities enabling them to utilize chitin as the sole source of carbon and nitrogen. Interestingly, several bacterial chitinases may also be involved in host pathogenesis. For example, in the important foodborne pathogen Listeria monocytogenes, the chitinases ChiA and ChiB and the lytic polysaccharide monooxygenase Lmo2467 are implicated in chitin assimilation but also act as virulence factors during the infection of mammalian hosts. Therefore, it is important to identify their regulators and induction cues to understand how the different roles of the chitinolytic system are controlled and mediated. Here, we provide evidence for the importance of lmo0327 and lmo0325, encoding a putative internalin/autolysin and a putative transcriptional activator, respectively, in the efficient expression of chitinase activity in L. monocytogenes and thereby provide new information regarding the function of the lmo0325-lmo0327 locus.
Highlights
The chitinolytic system of Listeria monocytogenes far comprises two chitinases, ChiA and ChiB, and a lytic polysaccharide monooxygenase, Lmo2467
We provide evidence for the importance of lmo0327 and lmo0325, encoding a putative internalin/autolysin and a putative transcriptional activator, respectively, in the efficient expression of chitinase activity in L. monocytogenes and thereby provide new information regarding the function of the lmo0325-lmo0327 locus
Lmo0327 Affects Listerial Chitinase Expression screening, as we have found it to be a strain of relatively high chitinolytic potential, a trait that can facilitate the identification of mutants
Summary
The chitinolytic system of Listeria monocytogenes far comprises two chitinases, ChiA and ChiB, and a lytic polysaccharide monooxygenase, Lmo2467. IMPORTANCE Many bacteria from terrestrial and marine environments express chitinase activities enabling them to utilize chitin as the sole source of carbon and nitrogen. In the important foodborne pathogen Listeria monocytogenes, the chitinases ChiA and ChiB and the lytic polysaccharide monooxygenase Lmo2467 are implicated in chitin assimilation and act as virulence factors during the infection of mammalian hosts. Chitin is an insoluble aminopolysaccharide comprising repeated alternating N-acetylglucosamine (GlcNAc) units and is present in abundance in marine and soil environments [1]. Central to the scavenging and catabolism of chitin are chitinases and associated chitin-binding proteins/lytic polysaccharide monooxygenases (LPMOs), which coordinately allow the bacteria to degrade the chitin to assimilative chitooligosaccharides, including the N,N=-diacetylchitobiose [(GlcNAc)2] dimer and the GlcNAc monomer. The components responsible for the recognition of chitin and subsequent induction of the chitinolytic system include a chitooligosaccharide-specific porin [23,24,25], encoded by chiP, as well as ChiS, a hybrid sensor kinase that is proposed to respond to chitooligosaccharides introduced into the periplasmic space through the action of ChiP and other porins [14]
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