Roles of Two Shewanella oneidensis MR-1 Extracellular Endonucleases

Department of Ecophysiology, Max-Planck-Institut fu¨r terrestrische Mikrobiologie, Marburg, Germany.
Applied and Environmental Microbiology (Impact Factor: 3.67). 06/2011; 77(15):5342-51. DOI: 10.1128/AEM.00643-11
Source: PubMed


The dissimilatory iron-reducing bacterium Shewanella oneidensis MR-1 is capable of using extracellular DNA (eDNA) as the sole source of carbon, phosphorus, and nitrogen. In addition, we
recently demonstrated that S. oneidensis MR-1 requires eDNA as a structural component during all stages of biofilm formation. In this study, we characterize the roles
of two Shewanella extracellular endonucleases, ExeS and ExeM. While ExeS is likely secreted into the medium, ExeM is predicted to remain associated
with the cell envelope. Both exeM and exeS are highly expressed under phosphate-limited conditions. Mutants lacking exeS and/or exeM exhibit decreased eDNA degradation; however, the capability of S. oneidensis MR-1 to use DNA as the sole source of phosphorus is only affected in mutants lacking exeM. Neither of the two endonucleases alleviates toxic effects of increased eDNA concentrations. The deletion of exeM and/or exeS significantly affects biofilm formation of S. oneidensis MR-1 under static conditions, and expression of exeM and exeS drastically increases during static biofilm formation. Under hydrodynamic conditions, a deletion of exeM leads to altered biofilms that consist of densely packed structures which are covered by a thick layer of eDNA. Based on
these results, we hypothesize that a major role of ExeS and, in particular, ExeM of S. oneidensis MR-1, is to degrade eDNA as a matrix component during biofilm formation to improve nutrient supply and to enable detachment.

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    • "Beyond its sheer abundance, eDNA is a major component of the microbial ecosystem as a dynamic reservoir of carbon (C), nitrogen (N), phosphorus (P), nucleotides, and genetic information (Dell'Anno and Danovaro, 2005; Corinaldesi et al., 2007, 2008). eDNA is engaged by prokaryotes through often complex endogenous mechanisms, including degradation by unbound and cellsurface-bound secreted nucleases (Heins et al., 1967; Provvedi et al., 2001; Sakamoto et al., 2001; Schmidt et al., 2007; Godeke et al., 2011a), import and export systems mediating natural DNA uptake (Chen and Dubnau, 2004; Maier et al., 2004; Chen et al., 2005; Averhoff, 2009), and as a major component of biofilms (Steinberger and Holden, 2005; Godeke et al., 2011b; Kiedrowski et al., 2011; Gloag et al., 2013). "
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    ABSTRACT: Extracellular DNA is found in all environments and is a dynamic component of the microbial ecosystem. Microbial cells produce and interact with extracellular DNA through many endogenous mechanisms. Extracellular DNA is processed and internalized for use as genetic information and as a major source of macronutrients, and plays several key roles within prokaryotic biofilms. Hypersaline sites contain some of the highest extracellular DNA concentrations measured in nature-a potential rich source of carbon, nitrogen, and phosphorus for halophilic microorganisms. We conducted DNA growth studies for the halophilic archaeon Haloferax volcanii DS2 and show that this model Halobacteriales strain is capable of using exogenous double-stranded DNA as a nutrient. Further experiments with varying medium composition, DNA concentration, and DNA types revealed that DNA is utilized primarily as a phosphorus source, that growth on DNA is concentration-dependent, and that DNA isolated from different sources is metabolized selectively, with a bias against highly divergent methylated DNA. Additionally, fluorescence microscopy showed that labeled DNA co-localized with H. volcanii cells. The gene Hvo_1477 was also identified using a comparative genomic approach as a factor likely to be involved in DNA processing at the cell surface, and deletion of Hvo_1477 created a strain deficient in the ability to grow on extracellular DNA. Widespread distribution of Hvo_1477 homologs in archaea suggests metabolism of extracellular DNA may be of broad ecological and physiological relevance in this domain of life.
    Full-text · Article · Feb 2014 · Frontiers in Microbiology
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    • "However, studies with isogenic mutants have provided evidence that extracellular nucleases keep the growth of biofilms in check. In Shewanella oneidensis, the disruption of genes encoding two extracellular nucleases, ExeS and ExeM, resulted in altered biofilm formation and the accumulation of eDNA (Godeke et al. 2011). Disruption of a third nuclease-encoding gene, endA, had little effect on biofilm structure (Heun et al. 2012). "
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    ABSTRACT: The death and lysis of microbial cells leads to the release of cytoplasmic contents, many of which are rapidly degraded by enzymes. However, some macromolecules survive intact and find new functions in the extracellular environment. There is now strong evidence that DNA released from cells during lysis, or sometimes by active secretion, becomes a key component of the macromolecular scaffold in many different biofilms. Enzymatic degradation of eDNA can weaken the biofilm structure and release microbial cells from the surface. Many bacteria produce extracellular deoxyribonuclease (DNase) enzymes that are apparently tightly regulated to avoid excessive degradation of the biofilm matrix. Interfering with these control mechanisms, or adding exogenous DNases, could prove a potent strategy for controlling biofilm growth. This article is protected by copyright. All rights reserved.
    Full-text · Article · Jul 2013 · Letters in Applied Microbiology
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    • "There is an alkaline phosphatase expressed upstream of the DNase, EddA, which may also be required for phosphorus acquisition from DNA. In Shewanella oneidensis, a secreted DNase (ExeM) with significant homology to EddB (34% identity) is also required for utilization of DNA as a nutrient source (Godeke et al., 2011). A number of intracellular ssDNA exonucleases have also been shown to be required for growth using DNA as a sole carbon course (Palchevskiy and Finkel, 2006). "
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    ABSTRACT: Extracellular DNA (eDNA) is in the environment, bodily fluids, in the matrix of biofilms, and accumulates at infection sites. eDNA can function as a nutrient source, a universal biofilm matrix component, and an innate immune effector in eDNA traps. In biofilms, eDNA is required for attachment, aggregation, and stabilization of microcolonies. We have recently shown that eDNA can sequester divalent metal cations, which has interesting implications on antibiotic resistance. eDNA binds metal cations and thus activates the Mg(2+)-responsive PhoPQ and PmrAB two-component systems. In Pseudomonas aeruginosa and many other Gram-negative bacteria, the PhoPQ/PmrAB systems control various genes required for virulence and resisting killing by antimicrobial peptides (APs), including the pmr genes (PA3552-PA3559) that are responsible for the addition of aminoarabinose to lipid A. The PA4773-PA4775 genes are a second DNA-induced cluster and are required for the production of spermidine on the outer surface, which protects the outer membrane from AP treatment. Both modifications mask the negative surface charges and limit membrane damage by APs. DNA-enriched biofilms or planktonic cultures have increased antibiotic resistance phenotypes to APs and aminoglycosides. These dual antibiotic resistance and immune evasion strategies may be expressed in DNA-rich environments and contribute to long-term survival.
    Full-text · Article · Feb 2013 · Frontiers in Microbiology
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