Type 3 Fimbriae and Biofilm Formation Are Regulated by the Transcriptional Regulators MrkHI in Klebsiella pneumoniae

Abstract

Klebsiella pneumoniae is an opportunistic pathogen which frequently causes hospital-acquired urinary and respiratory tract infections. K. pneumoniae may establish these infections in vivo following adherence, using the type 3 fimbriae, to indwelling devices coated with extracellular matrix components. Using a colony immunoblot screen, we identified transposon insertion mutants which were deficient for type 3 fimbrial surface production. One of these mutants possessed a transposon insertion within a gene, designated mrkI, encoding a putative transcriptional regulator. A site-directed mutant of this gene was constructed and shown to be deficient for fimbrial surface expression under aerobic conditions. MrkI mutants have a significantly decreased ability to form biofilms on both abiotic and extracellular matrix-coated surfaces. This gene was found to be cotranscribed with a gene predicted to encode a PilZ domain-containing protein, designated MrkH. This protein was found to bind cyclic-di-GMP (c-di-GMP) and regulate type 3 fimbrial expression.

Klebsiella pneumoniae yfiRNB operon affects biofilm formation, polysaccharide production and drug susceptibility

“…1a) has been previously implicated in biofilm formation in K. pneumoniae due to mutations that affect the synthesis of c-di-GMP, which is involved in the regulation of type 3 fimbriae expression (Johnson & Clegg, 2010;Johnson et al, 2011;Wilksch et al, 2011). Consistent with this, we identified a mutant via transposon mutagenesis of K pneumoniae clinical isolate CG217 (CG217 M39) (Suescú n et al, 2006) that mapped to the yfiR gene, which negatively controlled the YfiN DGC protein, and thus formed a biofilm more robust than that of the parental strain (Fig.…”

“…The cellular levels of c-di-GMP are modulated by diguanylate cyclases (DGCs) and phosphodiesterases, which catalyse its synthesis and degradation, respectively (Boyd & O'Toole, 2012;Römling et al, 2013). In K. pneumoniae, the c-di-GMP turnover enzymes YfiN and MrkJ have been shown to modulate type 3 fimbriae expression via the transcriptional regulator MrkH, which contains a c-di-GMP-binding PilZ domain (Johnson & Clegg, 2010;Johnson et al, 2011;Wilksch et al, 2011).…”

“…Biofilm formation in K. pneumoniae is affected by mutations in genes responsible for capsule biosynthesis and surface EPSs (Balestrino et al, 2008;Boddicker et al, 2006), genes for the synthesis of cell surface type 3 fimbriae (Di Martino et al, 2003;Langstraat et al, 2001), by mutations that affect quorum sensing (Balestrino et al, 2005) and by the intracellular levels of the second messenger c-di-GMP (Johnson & Clegg, 2010;Johnson et al, 2011;Wilksch et al, 2011). The cellular levels of c-di-GMP are modulated by diguanylate cyclases (DGCs) and phosphodiesterases, which catalyse its synthesis and degradation, respectively (Boyd & O'Toole, 2012;Römling et al, 2013).…”

FimK regulation on the expression of type 1 fimbriae in Klebsiella pneumoniae CG43S3

“…As shown in Fig. 5(a), the purified recombinant FimK exhibited considerably less bis-PNPP catalytic activity than the recombinant MrkJ, for which PDE activity has been reported (Johnson et al, 2011). Recombinant clones expressing EIL fimK and AIL fimK containing the C-terminal EIL domain of FimK and the domain with critical E245 residue replaced with alanine, respectively, were also generated and the corresponding overexpressed proteins were purified.…”

Structures of the activator of K. pneumonia biofilm formation, MrkH, indicates PilZ domains involved in c-di-GMP and DNA binding

“…The pathogenicity of K. pneumonia is associated with its ability to form biofilms, which are largely recalcitrant to treatment (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11)(12)(13)(14). Key to K. pneumonia biofilm development are mannose-resistant Klebsiella-like (Mrk) hemagglutinins or Mrk proteins, which are encoded by the mrkABCDF operon (7)(8)(9)(10)(11)(12)(13)(14).…”