The Disulfide Bonding System Suppresses CsgD-Independent Cellulose Production in Escherichia coli

Hufnagel, David A.; DePas, William H.; Chapman, Matthew R.

doi:10.1128/jb.02019-14

Cited by 31 publications

(59 citation statements)

References 77 publications

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“…2A; see also Fig. S2 in the supplemental material) (19). We hypothesize this is due to increased metabolic activity as nutrient limitation was previously shown to increase CsgD activity and biofilm formation (23).…”

Section: Resultsmentioning

confidence: 99%

“…Western blotting was performed as previously described (19,21). Rugose biofilms were collected with blue inoculating loops and resuspended in 1 ml 50 mM potassium phosphate (KPi) buffer (pH 7.2) before being vortexed briefly.…”

Section: Methodsmentioning

confidence: 99%

“…Miller assays were performed as previously described (19,22). Rugose biofilms were resuspended in KPi buffer and diluted 1:10.…”

Section: Methodsmentioning

confidence: 99%

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The Catabolite Repressor Protein-Cyclic AMP Complex Regulates csgD and Biofilm Formation in Uropathogenic Escherichia coli

et al. 2016

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The extracellular matrix protects Escherichia coli from immune cells, oxidative stress, predation, and other environmental stresses. Production of the E. coli extracellular matrix is regulated by transcription factors that are tuned to environmental conditions. The biofilm master regulator protein CsgD upregulates curli and cellulose, the two major polymers in the extracellular matrix of uropathogenic E. coli (UPEC) biofilms. We found that cyclic AMP (cAMP) regulates curli, cellulose, and UPEC biofilms through csgD. The alarmone cAMP is produced by adenylate cyclase (CyaA), and deletion of cyaA resulted in reduced extracellular matrix production and biofilm formation. The catabolite repressor protein (CRP) positively regulated csgD transcription, leading to curli and cellulose production in the UPEC isolate, UTI89. Glucose, a known inhibitor of CyaA activity, blocked extracellular matrix formation when added to the growth medium. The mutant strains ⌬cyaA and ⌬crp did not produce rugose biofilms, pellicles, curli, cellulose, or CsgD. Three putative CRP binding sites were identified within the csgD-csgB intergenic region, and purified CRP could gel shift the csgD-csgB intergenic region. Additionally, we found that CRP binded upstream of kpsMT, which encodes machinery for K1 capsule production. Together our work shows that cAMP and CRP influence E. coli biofilms through transcriptional regulation of csgD. IMPORTANCE The catabolite repressor protein (CRP)-cyclic AMP (cAMP) complex influences the transcription of ϳ7% of genes on the Escherichia coli is a multifaceted Gram-negative bacterium that is equipped for survival in disparate environments ranging from the gastrointestinal and urinary tracts to waterways and soil (1). Growth of E. coli can occur with either planktonic or biofilm lifestyles, the transition between which requires a highly organized shift in gene expression. The biofilm lifestyle is characterized by adherence, slow growth, resistance to environmental insults, and production of an extracellular matrix (ECM) (2). CsgD is the major biofilm regulator of E. coli and of many other bacteria of the Enterobacteriaceae family (3-5), and it promotes the production of two E. coli surface appendages, the amyloid fiber curli and the fibrous glycan chain cellulose (2). CsgD represses the transcription of motility-associated genes, while mediating self and surface attachment (6-8).E. coli is found in ϳ80% of urinary tract infections (UTI), more than any other causative agent, and has the ability to thrive in the bladder and urinary tract environments (9). During UTI, uropathogenic E. coli (UPEC) uses various means to evade and colonize the host. Curli and other factors function in establishing an initial bacterial titer (10). Once in the bladder, E. coli utilizes type 1 pili to invade bladder epithelial cells (11). E. coli bacteria then form a biofilm-like structure known as an intracellular bacterial community (IBC), and cells are coated by the ECM component K1 for protection from host cell recognition ...

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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The Catabolite Repressor Protein-Cyclic AMP Complex Regulates csgD and Biofilm Formation in Uropathogenic Escherichia coli

et al. 2016

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“…Mutation of yfiR affects swarming and swimming motility, as well as early biofilm formation (28,30). Furthermore, the Yfi system promotes CsgD-independent cellulose production under reducing conditions (31). Likewise, in the UPEC strain CFT073, yfiR mutants are affected in synthesis of curli and cellulose, and they are attenuated in mouse bladder and kidney infection (32).…”

Section: Resultsmentioning

confidence: 99%

YjjQ Represses Transcription of flhDC and Additional Loci in Escherichia coli

et al. 2015

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The presumptive transcriptional regulator YjjQ has been identified as being virulence associated in avian pathogenic Escherichia coli (APEC). In this work, we characterize YjjQ as transcriptional repressor of the flhDC operon, encoding the master regulator of flagellar synthesis, and of additional loci. The latter include gfc (capsule 4 synthesis), ompC (outer membrane porin C), yfiRNB (regulated c-di-GMP synthesis), and loci of poorly defined function (ybhL and ymiA-yciX). We identify the YjjQ DNA-binding sites at the flhDC and gfc promoters and characterize a DNA-binding sequence motif present at all promoters found to be repressed by YjjQ. At the flhDC promoter, the YjjQ DNA-binding site overlaps the RcsA-RcsB DNA-binding site. RcsA-RcsB likewise represses the flhDC promoter, but the repression by YjjQ and that by RcsA-RcsB are independent of each other. These data suggest that YjjQ is an additional regulator involved in the complex control of flhDC at the level of transcription initiation. Furthermore, we show that YjjQ represses motility of the E. coli K-12 laboratory strain and of uropathogenic E. coli (UPEC) strains CFT073 and 536. Regulation of flhDC, yfiRNB, and additional loci by YjjQ may be features relevant for pathogenicity. IMPORTANCEEscherichia coli is a commensal and pathogenic bacterium causing intra-and extraintestinal infections in humans and farm animals. The pathogenicity of E. coli strains is determined by their particular genome content, which includes essential and associated virulence factors that control the cellular physiology in the host environment. However, the gene pools of commensal and pathogenic E. coli are not clearly differentiated, and the function of virulence-associated loci needs to be characterized. In this study, we characterize the function of yjjQ, encoding a transcription regulator that was identified as being virulence associated in avian pathogenic E. coli (APEC). We characterize YjjQ as transcriptional repressor of flagellar motility and of additional loci related to pathogenicity.

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“…With some rare exceptions (27)(28)(29)(30), analyses of GGDEF/EAL domain genes or proteins of E. coli have been performed with laboratory K-12 strains. Therefore, almost nothing is known about how the complement of GGDEF/EAL domain-encoding genes may vary in different E. coli strains, in particular when comparing pathogenic and commensal strains.…”

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Genome-Based Comparison of Cyclic Di-GMP Signaling in Pathogenic and Commensal Escherichia coli Strains

Povolotsky

Hengge

2016

J Bacteriol

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The ubiquitous bacterial second messenger cyclic di-GMP (c-di-GMP) has recently become prominent as a trigger for biofilm formation in many bacteria. It is generated by diguanylate cyclases (DGCs; with GGDEF domains) and degraded by specific phosphodiesterases (PDEs; containing either EAL or HD-GYP domains). Most bacterial species contain multiples of these proteins with some having specific functions that are based on direct molecular interactions in addition to their enzymatic activities. Escherichia coli K-12 laboratory strains feature 29 genes encoding GGDEF and/or EAL domains, resulting in a set of 12 DGCs, 13 PDEs, and four enzymatically inactive "degenerate" proteins that act by direct macromolecular interactions. We present here a comparative analysis of GGDEF/EAL domain-encoding genes in 61 genomes of pathogenic, commensal, and probiotic E. coli strains (including enteric pathogens such as enteroaggregative, enterohemorrhagic, enteropathogenic, enterotoxigenic, and adherent and invasive Escherichia coli and the 2011 German outbreak O104:H4 strain, as well as extraintestinal pathogenic E. coli, such as uropathogenic and meningitis-associated E. coli). We describe additional genes for two membrane-associated DGCs (DgcX and DgcY) and four PDEs (the membrane-associated PdeT, as well as the EAL domain-only proteins PdeW, PdeX, and PdeY), thus showing the pangenome of E. coli to contain at least 35 GGDEF/EAL domain proteins. A core set of only eight proteins is absolutely conserved in all 61 strains: DgcC (YaiC), DgcI (YliF), PdeB (YlaB), PdeH (YhjH), PdeK (YhjK), PdeN (Rtn), and the degenerate proteins CsrD and CdgI (YeaI). In all other GGDEF/EAL domain genes, diverse point and frameshift mutations, as well as small or large deletions, were discovered in various strains. IMPORTANCEOur analysis reveals interesting trends in pathogenic Escherichia coli that could reflect different host cell adherence mechanisms. These may either benefit from or be counteracted by the c-di-GMP-stimulated production of amyloid curli fibers and cellulose. Thus, EAEC, which adhere in a "stacked brick" biofilm mode, have a potential for high c-di-GMP accumulation due to DgcX, a strongly expressed additional DGC. In contrast, EHEC and UPEC, which use alternative adherence mechanisms, tend to have extra PDEs, suggesting that low cellular c-di-GMP levels are crucial for these strains under specific conditions. Overall, our study also indicates that GGDEF/EAL domain proteins evolve rapidly and thereby contribute to adaptation to host-specific and environmental niches of various types of E. coli.

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The Disulfide Bonding System Suppresses CsgD-Independent Cellulose Production in Escherichia coli

Cited by 31 publications

References 77 publications

The Catabolite Repressor Protein-Cyclic AMP Complex Regulates csgD and Biofilm Formation in Uropathogenic Escherichia coli

The Catabolite Repressor Protein-Cyclic AMP Complex Regulates csgD and Biofilm Formation in Uropathogenic Escherichia coli

YjjQ Represses Transcription of flhDC and Additional Loci in Escherichia coli

Genome-Based Comparison of Cyclic Di-GMP Signaling in Pathogenic and Commensal Escherichia coli Strains

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