An official website of the United States government
Abstract Glycerol dibiphytanyl glycerol tetraethers (GDGTs) are archaeal monolayer membrane lipids that can provide a competitive advantage in extreme environments. Here, we identify a radical SAM protein, tetraether synthase (Tes), that participates in the synthesis of GDGTs. Attempts to generate a tes-deleted mutant in Sulfolobus acidocaldarius were unsuccessful, suggesting that the gene is essential in this organism. Heterologous expression of tes homologues leads to production of GDGT and structurally related lipids in the methanogen Methanococcus maripaludis (which otherwise does not synthesize GDGTs and lacks a tes homolog, but produces a putative GDGT precursor, archaeol). Tes homologues are encoded in the genomes of many archaea, as well as in some bacteria, in which they might be involved in the synthesis of bacterial branched glycerol dialkyl glycerol tetraethers.
more »
« less
Stay or Go: Sulfolobales Biofilm Dispersal Is Dependent on a Bifunctional VapB Antitoxin
ABSTRACT A type II VapB14 antitoxin regulates biofilm dispersal in the archaeal thermoacidophile Sulfolobus acidocaldarius through traditional toxin neutralization but also through noncanonical transcriptional regulation. Type II VapC toxins are ribonucleases that are neutralized by their proteinaceous cognate type II VapB antitoxin. VapB antitoxins have a flexible tail at their C terminus that covers the toxin’s active site, neutralizing its activity. VapB antitoxins also have a DNA-binding domain at their N terminus that allows them to autorepress not only their own promoters but also distal targets. VapB14 antitoxin gene deletion in S. acidocaldarius stunted biofilm and planktonic growth and increased motility structures (archaella). Conversely, planktonic cells were devoid of archaella in the Δ vapC14 cognate toxin mutant. VapB14 is highly conserved at both the nucleotide and amino acid levels across the Sulfolobales, extremely unusual for type II antitoxins, which are typically acquired through horizontal gene transfer. Furthermore, homologs of VapB14 are found across the Crenarchaeota , in some Euryarchaeota , and even bacteria. S. acidocaldarius vapB14 and its homolog in the thermoacidophile Metallosphaera sedula (Msed_0871) were both upregulated in biofilm cells, supporting the role of the antitoxin in biofilm regulation. In several Sulfolobales species, including M. sedula, homologs of vapB14 and vapC14 are not colocalized. Strikingly, Sulfuracidifex tepidarius has an unpaired VapB14 homolog and lacks a cognate VapC14, illustrating the toxin-independent conservation of the VapB14 antitoxin. The findings here suggest that a stand-alone VapB-type antitoxin was the product of selective evolutionary pressure to influence biofilm formation in these archaea, a vital microbial community behavior. IMPORTANCE Biofilms allow microbes to resist a multitude of stresses and stay proximate to vital nutrients. The mechanisms of entering and leaving a biofilm are highly regulated to ensure microbial survival, but are not yet well described in archaea. Here, a VapBC type II toxin-antitoxin system in the thermoacidophilic archaeon Sulfolobus acidocaldarius was shown to control biofilm dispersal through a multifaceted regulation of the archaeal motility structure, the archaellum. The VapC14 toxin degrades an RNA that causes an increase in archaella and swimming. The VapB14 antitoxin decreases archaella and biofilm dispersal by binding the VapC14 toxin and neutralizing its activity, while also repressing the archaellum genes. VapB14-like antitoxins are highly conserved across the Sulfolobales and respond similarly to biofilm growth. In fact, VapB14-like antitoxins are also found in other archaea, and even in bacteria, indicating an evolutionary pressure to maintain this protein and its role in biofilm formation.
more »
« less
- Award ID(s):
- 1802939
- PAR ID:
- 10423922
- Editor(s):
- Sauer, Karin; Lee, Sang Yup
- Date Published:
- Journal Name:
- mBio
- Volume:
- 14
- Issue:
- 2
- ISSN:
- 2150-7511
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
More Like this
-
-
Summary Microorganisms regulate the composition of their membranes in response to environmental cues. Many Archaea maintain the fluidity and permeability of their membranes by adjusting the number of cyclic moieties within the cores of their glycerol dibiphytanyl glycerol tetraether (GDGT) lipids. Cyclized GDGTs increase membrane packing and stability, which has been shown to help cells survive shifts in temperature and pH. However, the extent of this cyclization also varies with growth phase and electron acceptor or donor limitation. These observations indicate a relationship between energy metabolism and membrane composition. Here we show that the average degree of GDGT cyclization increases with doubling time in continuous cultures of the thermoacidophileSulfolobus acidocaldarius(DSM 639). This is consistent with the behavior of a mesoneutrophile,Nitrosopumilus maritimusSCM1. Together, these results demonstrate that archaeal GDGT distributions can shift in response to electron donor flux and energy availability, independent of pH or temperature. Paleoenvironmental reconstructions based on GDGTs thus capture the energy available to microbes, which encompasses fluctuations in temperature and pH, as well as electron donor and acceptor availability. The ability of Archaea to adjust membrane composition and packing may be an important strategy that enables survival during episodes of energy stress.more » « less
-
Abstract Lipoproteins are major constituents of prokaryotic cell surfaces. In bacteria, lipoprotein attachment to membrane lipids is catalyzed by prolipoprotein diacylglyceryl transferase (Lgt). However, no Lgt homologs have been identified in archaea, suggesting the unique archaeal membrane lipids require distinct enzymes for lipoprotein lipidation. Here, we performedin silicopredictions for all major archaeal lineages and revealed a high prevalence of lipoproteins across the domain Archaea. Using comparative genomics, we identified the first set of candidates for archaeal lipoprotein biogenesis components (Ali). Genetic and biochemical characterization confirmed two paralogous genes,aliAandaliB, are important for lipoprotein lipidation in the archaeonHaloferax volcanii. Disruption of AliA- and AliB-mediated lipoprotein lipidation results in severe growth defects, decreased motility, and cell-shape alterations, underscoring the importance of lipoproteins in archaeal cell physiology. AliA and AliB also exhibit different enzymatic activities, including potential substrate selectivity, uncovering a new layer of regulation for prokaryotic lipoprotein lipidation.more » « less
-
Svensson, Sarah L (Ed.)Bacterial chromosomal type I toxin-antitoxin systems consist of a small protein, typically under 60 amino acids, and a small RNA (sRNA) that represses toxin translation. These gene pairs have gained attention over the last decade for their contribution to antibiotic persistence and phage tolerance in bacteria. However, biological functions for many remain elusive as gene deletions often fail to produce an observable phenotype. For many pairs, it is still unknown when the toxin and/or antitoxin gene are natively expressed within the bacterium. We examined sequence conservation of three type I toxin-antitoxin systems,tisB/istR-1, shoB/ohsC, and zor/orz, in over 2,000Escherichia colistrains, including pathogenic and commensal isolates. Using our custom database, we found that these gene pairs are widespread acrossE. coliand have expression potential via BLASTn. We identified an alternative, dominant sequence variant of TisB and confirmed that it is toxic upon overproduction. Additionally, analyses revealed a highly conserved sequence in thezorOmRNA untranslated region that is required for full toxicity. We further noted that over 30% ofE. coligenomes contain anorzantitoxin gene only and confirmed its expression in a representative strain: the first confirmed report of a type I antitoxin without its cognate toxin. Our results add to our understanding of these systems, and our methodology is applicable for other type I loci to identify critical regulatory and functional features.IMPORTANCEChromosomal type I toxin-antitoxins are a class of genes that have gained increasing attention over the last decade for their roles in antibiotic persistence which may contribute to therapeutic failures. However, the control of many of these genes and when they function have remained elusive. We demonstrate that a simple genetic conservation-based approach utilizing free, publicly available data yields known and novel insights into the regulation and function of three chromosomal type I toxin-antitoxins inEscherichia coli. This study also provides a framework for how this approach could be applied to other genes of interest.more » « less
-
Maupin-Furlow, Julie A (Ed.)ABSTRACT Many prokaryotes use swimming motility to move toward favorable conditions and escape adverse surroundings. Regulatory mechanisms governing bacterial flagella-driven motility are well-established; however, little is yet known about the regulation underlying swimming motility propelled by the archaeal cell surface structure, the archaella. Previous research showed that the deletion of the adhesion pilins (PilA1-6), subunits of the type IV pili cell surface structure, renders the model archaeonHaloferax volcaniinon-motile. In this study, we used ethyl methanesulfonate mutagenesis and a motility assay to identify motile suppressors of the ∆pilA[1-6] strain. Of the eight suppressors identified, six contain missense mutations in archaella biosynthesis genes,arlIandarlJ. In transexpression ofarlIandarlJmutant constructs in the respective multi-deletion strains ∆pilA[1-6]∆arlIand ∆pilA[1-6]∆arlJconfirmed their role in suppressing the ∆pilA[1-6] motility defect. Additionally, three suppressors harbor co-occurring disruptive missense and nonsense mutations incirA, a gene encoding a proposed regulatory protein. A deletion ofcirAresulted in hypermotility, whilecirAexpressionin transin wild-type cells led to decreased motility. Moreover, quantitative real-time PCR analysis revealed that in wild-type cells, higher expression levels ofarlI,arlJ, and the archaellin genearlA1were observed in motile early-log phase rod-shaped cells compared to non-motile mid-log phase disk-shaped cells. Conversely, ∆cirAcells, which form rods during both early- and mid-log phases, exhibited similar expression levels ofarlgenes in both growth phases. Our findings contribute to a deeper understanding of the mechanisms governing archaeal motility, highlighting the involvement of ArlI, ArlJ, and CirA in pilin-mediated motility regulation.IMPORTANCEArchaea are close relatives of eukaryotes and play crucial ecological roles. Certain behaviors, such as swimming motility, are thought to be important for archaeal environmental adaptation. Archaella, the archaeal motility appendages, are evolutionarily distinct from bacterial flagella, and the regulatory mechanisms driving archaeal motility are largely unknown. Previous research has linked the loss of type IV pili subunits to archaeal motility suppression. This study reveals threeHaloferax volcaniiproteins involved in pilin-mediated motility regulation, offering a deeper understanding of motility regulation in this understudied domain while also paving the way for uncovering novel mechanisms that govern archaeal motility. Understanding archaeal cellular processes will help elucidate the ecological roles of archaea as well as the evolution of these processes across domains.more » « less
- Free Publicly Accessible Full Text
-
Accepted Manuscript1.0
- Journal Article:
- https://doi.org/10.1128/mbio.00053-23
