More Like this

1. Validating the Cyc2 Neutrophilic Iron Oxidation Pathway Using Meta-omics of Zetaproteobacteria Iron Mats at Marine Hydrothermal Vents

   https://doi.org/10.1128/mSystems.00553-19  

   McAllister, Sean M.; Polson, Shawn W.; Butterfield, David A.; Glazer, Brian T.; Sylvan, Jason B.; Chan, Clara S.; Lloyd, Karen G. (February 2020, mSystems)

   ABSTRACT Zetaproteobacteria create extensive iron (Fe) oxide mats at marine hydrothermal vents, making them an ideal model for microbial Fe oxidation at circumneutral pH. Comparison of neutrophilic Fe oxidizer isolate genomes has revealed a hypothetical Fe oxidation pathway, featuring a homolog of the Fe oxidase Cyc2 from Acidithiobacillus ferrooxidans . However, Cyc2 function is not well verified in neutrophilic Fe oxidizers, particularly in Fe-oxidizing environments. Toward this, we analyzed genomes and metatranscriptomes of Zetaproteobacteria , using 53 new high-quality metagenome-assembled genomes reconstructed from Fe mats at Mid-Atlantic Ridge, Mariana Backarc, and Loihi Seamount (Hawaii) hydrothermal vents. Phylogenetic analysis demonstrated conservation of Cyc2 sequences among most neutrophilic Fe oxidizers, suggesting a common function. We confirmed the widespread distribution of cyc2 and other model Fe oxidation pathway genes across all represented Zetaproteobacteria lineages. High expression of these genes was observed in diverse Zetaproteobacteria under multiple environmental conditions and in incubations. The putative Fe oxidase gene cyc2 was highly expressed in situ , often as the top expressed gene. The cyc2 gene showed increased expression in Fe(II)-amended incubations, with corresponding increases in carbon fixation and central metabolism gene expression. These results substantiate the Cyc2-based Fe oxidation pathway in neutrophiles and demonstrate its significance in marine Fe-mineralizing environments. IMPORTANCE Iron oxides are important components of our soil, water supplies, and ecosystems, as they sequester nutrients, carbon, and metals. Microorganisms can form iron oxides, but it is unclear whether this is a significant mechanism in the environment. Unlike other major microbial energy metabolisms, there is no marker gene for iron oxidation, hindering our ability to track these microbes. Here, we investigate a promising possible iron oxidation gene, cyc2 , in iron-rich hydrothermal vents, where iron-oxidizing microbes dominate. We pieced together diverse Zetaproteobacteria genomes, compared these genomes, and analyzed expression of cyc2 and other hypothetical iron oxidation genes. We show that cyc2 is widespread among iron oxidizers and is highly expressed and potentially regulated, making it a good marker for the capacity for iron oxidation and potentially a marker for activity. These findings will help us understand and potentially quantify the impacts of neutrophilic iron oxidizers in a wide variety of marine and terrestrial environments. 

   more » « less
2. Critical assessment of pan-genomic analysis of metagenome-assembled genomes

   https://doi.org/10.1093/bib/bbac413  

   Li, Tang; Yin, Yanbin (September 2022, Briefings in Bioinformatics)

   Abstract Pan-genome analyses of metagenome-assembled genomes (MAGs) may suffer from the known issues with MAGs: fragmentation, incompleteness and contamination. Here, we conducted a critical assessment of pan-genomics of MAGs, by comparing pan-genome analysis results of complete bacterial genomes and simulated MAGs. We found that incompleteness led to significant core gene (CG) loss. The CG loss remained when using different pan-genome analysis tools (Roary, BPGA, Anvi’o) and when using a mixture of MAGs and complete genomes. Contamination had little effect on core genome size (except for Roary due to in its gene clustering issue) but had major influence on accessory genomes. Importantly, the CG loss was partially alleviated by lowering the CG threshold and using gene prediction algorithms that consider fragmented genes, but to a less degree when incompleteness was higher than 5%. The CG loss also led to incorrect pan-genome functional predictions and inaccurate phylogenetic trees. Our main findings were supported by a study of real MAG-isolate genome data. We conclude that lowering CG threshold and predicting genes in metagenome mode (as Anvi’o does with Prodigal) are necessary in pan-genome analysis of MAGs. Development of new pan-genome analysis tools specifically for MAGs are needed in future studies. 

   more » « less
3. Regulation of Biofilm Exopolysaccharide Production by Cyclic Di-Guanosine Monophosphate

   https://doi.org/10.3389/fmicb.2021.730980  

   Poulin, Myles B.; Kuperman, Laura L. (September 2021, Frontiers in Microbiology)

   Many bacterial species in nature possess the ability to transition into a sessile lifestyle and aggregate into cohesive colonies, known as biofilms. Within a biofilm, bacterial cells are encapsulated within an extracellular polymeric substance (EPS) comprised of polysaccharides, proteins, nucleic acids, lipids, and other small molecules. The transition from planktonic growth to the biofilm lifecycle provides numerous benefits to bacteria, such as facilitating adherence to abiotic surfaces, evasion of a host immune system, and resistance to common antibiotics. As a result, biofilm-forming bacteria contribute to 65% of infections in humans, and substantially increase the energy and time required for treatment and recovery. Several biofilm specific exopolysaccharides, including cellulose, alginate, Pel polysaccharide, and poly- N -acetylglucosamine (PNAG), have been shown to play an important role in bacterial biofilm formation and their production is strongly correlated with pathogenicity and virulence. In many bacteria the biosynthetic machineries required for assembly of these exopolysaccharides are regulated by common signaling molecules, with the second messenger cyclic di-guanosine monophosphate (c - di-GMP) playing an especially important role in the post-translational activation of exopolysaccharide biosynthesis. Research on treatments of antibiotic-resistant and biofilm-forming bacteria through direct targeting of c-di-GMP signaling has shown promise, including peptide-based treatments that sequester intracellular c-di-GMP. In this review, we will examine the direct role c-di-GMP plays in the biosynthesis and export of biofilm exopolysaccharides with a focus on the mechanism of post-translational activation of these pathways, as well as describe novel approaches to inhibit biofilm formation through direct targeting of c-di-GMP. 

   more » « less
4. Differential ligand-selective control of opposing enzymatic activities within a bifunctional c-di-GMP enzyme

   https://doi.org/10.1073/pnas.2100657118  

   Patterson, Dayna C.; Ruiz, Myrrh Perez; Yoon, Hyerin; Walker, Johnnie A.; Armache, Jean-Paul; Yennawar, Neela H.; Weinert, Emily E. (September 2021, Proceedings of the National Academy of Sciences)

   Cyclic dimeric guanosine monophosphate (c-di-GMP) serves as a second messenger that modulates bacterial cellular processes, including biofilm formation. While proteins containing both c-di-GMP synthesizing (GGDEF) and c-di-GMP hydrolyzing (EAL) domains are widely predicted in bacterial genomes, it is poorly understood how domains with opposing enzymatic activity are regulated within a single polypeptide. Herein, we report the characterization of a globin-coupled sensor protein (GCS) fromPaenibacillus dendritiformis(DcpG) with bifunctional c-di-GMP enzymatic activity. DcpG contains a regulatory sensor globin domain linked to diguanylate cyclase (GGDEF) and phosphodiesterase (EAL) domains that are differentially regulated by gas binding to the heme; GGDEF domain activity is activated by the Fe(II)-NO state of the globin domain, while EAL domain activity is activated by the Fe(II)-O₂state. The in vitro activity of DcpG is mimicked in vivo by the biofilm formation ofP. dendritiformisin response to gaseous environment, with nitric oxide conditions leading to the greatest amount of biofilm formation. The ability of DcpG to differentially control GGDEF and EAL domain activity in response to ligand binding is likely due to the unusual properties of the globin domain, including rapid ligand dissociation rates and high midpoint potentials. Using structural information from small-angle X-ray scattering and negative stain electron microscopy studies, we developed a structural model of DcpG, providing information about the regulatory mechanism. These studies provide information about full-length GCS protein architecture and insight into the mechanism by which a single regulatory domain can selectively control output domains with opposing enzymatic activities. 

   more » « less
5. Quantitative input–output dynamics of a c-di-GMP signal transduction cascade in Vibrio cholerae

   https://doi.org/10.1371/journal.pbio.3001585  

   Bridges, Andrew A.; Prentice, Jojo A.; Fei, Chenyi; Wingreen, Ned S.; Bassler, Bonnie L. (March 2022, PLOS Biology)

   Waldor, Matthew K. (Ed.)

   Bacterial biofilms are multicellular communities that collectively overcome environmental threats and clinical treatments. To regulate the biofilm lifecycle, bacteria commonly transduce sensory information via the second messenger molecule cyclic diguanylate (c-di-GMP). Using experimental and modeling approaches, we quantitatively capture c-di-GMP signal transmission via the bifunctional polyamine receptor NspS-MbaA, from ligand binding to output, in the pathogen Vibrio cholerae . Upon binding of norspermidine or spermidine, NspS-MbaA synthesizes or degrades c-di-GMP, respectively, which, in turn, drives alterations specifically to biofilm gene expression. A long-standing question is how output specificity is achieved via c-di-GMP, a diffusible molecule that regulates dozens of effectors. We show that NspS-MbaA signals locally to specific effectors, sensitizing V . cholerae to polyamines. However, local signaling is not required for specificity, as changes to global cytoplasmic c-di-GMP levels can selectively regulate biofilm genes. This work establishes the input–output dynamics underlying c-di-GMP signaling, which could be useful for developing bacterial manipulation strategies. 

   more » « less