Putative Nucleotide-Based Second Messengers in the Archaeal Model Organisms Haloferax volcanii and Sulfolobus acidocaldarius

Research on nucleotide-based second messengers began in 1956 with the discovery of cyclic adenosine monophosphate (3′,5′-cAMP) by Earl Wilbur Sutherland and his co-workers. Since then, a broad variety of different signaling molecules composed of nucleotides has been discovered. These molecules fulfill crucial tasks in the context of intracellular signal transduction. The vast majority of the currently available knowledge about nucleotide-based second messengers originates from model organisms belonging either to the domain of eukaryotes or to the domain of bacteria, while the archaeal domain is significantly underrepresented in the field of nucleotide-based second messenger research. For several well-stablished eukaryotic and/or bacterial nucleotide-based second messengers, it is currently not clear whether these signaling molecules are present in archaea. In order to shed some light on this issue, this study analyzed cell extracts of two major archaeal model organisms, the euryarchaeon Haloferax volcanii and the crenarchaeon Sulfolobus acidocaldarius, using a modern mass spectrometry method to detect a broad variety of currently known nucleotide-based second messengers. The nucleotides 3′,5′-cAMP, cyclic guanosine monophosphate (3′,5′-cGMP), 5′-phosphoadenylyl-3′,5′-adenosine (5′-pApA), diadenosine tetraphosphate (Ap4A) as well as the 2′,3′-cyclic isomers of all four RNA building blocks (2′,3′-cNMPs) were present in both species. In addition, H. volcanii cell extracts also contain cyclic cytosine monophosphate (3′,5′-cCMP), cyclic uridine monophosphate (3′,5′-cUMP) and cyclic diadenosine monophosphate (3′,5′-c-di-AMP). The widely distributed bacterial second messengers cyclic diguanosine monophosphate (3′,5′-c-di-GMP) and guanosine (penta-)/tetraphosphate [(p)ppGpp] could not be detected. In summary, this study gives a comprehensive overview on the presence of a large set of currently established or putative nucleotide-based second messengers in an eury- and a crenarchaeal model organism.

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Biodegradation of the xenobiotic organic disulphide 4,4′-dithiodibutyric acid by Rhodococcus erythropolis strain MI2 and comparison with the microbial utilization of 3,3′-dithiodipropionic acid and 3,3′-thiodipropionic acid

Microbiology ◽

10.1099/mic.0.036178-0 ◽

2010 ◽

Vol 156 (4) ◽

pp. 1221-1233 ◽

Cited By ~ 10

Author(s):

Jan Hendrik Wübbeler ◽

Nadine Bruland ◽

Milena Wozniczka ◽

Alexander Steinbüchel

Keyword(s):

Carbon Source ◽

Succinic Acid ◽

Hydrogen Sulphide ◽

Rhodococcus Erythropolis ◽

Building Blocks ◽

Bacterial Strains ◽

Sulphur Compounds ◽

Promising Alternative ◽

Microbial Utilization ◽

Broad Variety

Application of the non-toxic 3,3′-thiodipropionic acid (TDP) and 3,3′-dithiodipropionic acid (DTDP) as precursors for the microbial production of polythioesters (PTEs), a class of biologically persistent biopolymers containing sulphur in the backbone, was successfully established previously. However, synthesis of PTEs containing 4-mercaptobutyrate (4MB) as building blocks could not be achieved. The very harmful 4MB is not used as a PTE precursor or as the carbon source for growth by any known strain. As a promising alternative, the harmless oxidized disulfide of two molecules of 4MB, 4,4′-dithiodibutyric acid (DTDB), was employed for enrichments of bacterial strains capable of biodegradation. Investigation of novel precursor substrates for PTEs and comparison of respective strains growing on TDP, DTDP and DTDB as sole carbon source was accomplished. A broad variety of bacteria capable of using one of these organic sulphur compounds were isolated and compared. TDP and DTDP were degraded by several strains belonging to different genera, whereas all DTDB-utilizing strains were affiliated to the species Rhodococcus erythropolis. Transposon mutagenesis of R. erythropolis strain MI2 and screening of 7500 resulting mutants yielded three mutants exhibiting impaired growth on DTDB. Physiological studies revealed production of volatile hydrogen sulphide and accumulation of significant amounts of 4MB, 4-oxo-4-sulphanylbutanoic acid and succinic acid in the culture supernatants. Based on this knowledge, a putative pathway for degradation of DTDB was proposed: DTDB could be cleaved into two molecules of 4MB, followed by an oxidation yielding 4-oxo-4-sulphanylbutanoic acid. A putative desulphydrase probably catalyses the abstraction of sulphur, thereby generating succinic acid and hydrogen sulphide.

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Hot and sweet: protein glycosylation in Crenarchaeota

Biochemical Society Transactions ◽

10.1042/bst20120296 ◽

2013 ◽

Vol 41 (1) ◽

pp. 384-392 ◽

Cited By ~ 33

Author(s):

Benjamin H. Meyer ◽

Sonja-Verena Albers

Keyword(s):

Living Cell ◽

Protein Glycosylation ◽

Haloferax Volcanii ◽

Present Review ◽

Sulfolobus Acidocaldarius ◽

Glycosylphosphatidylinositol Anchor ◽

Sweet Protein ◽

Domains Of Life ◽

Glycosylation Pathway ◽

Sugar Chains

Every living cell is covered with a dense and complex array of covalently attached sugars or sugar chains. The majority of these glycans are linked to proteins via the so-called glycosylation process. Protein glycosylation is found in all three domains of life: Eukarya, Bacteria and Archaea. However, on the basis of the limit in analytic tools for glycobiology and genetics in Archaea, only in the last few years has research on archaeal glycosylation pathways started mainly in the Euryarchaeota Haloferax volcanii, Methanocaldococcus maripaludis and Methanococcus voltae. Recently, major steps of the crenarchaeal glycosylation process of the thermoacidophilic archaeon Sulfolobus acidocaldarius have been described. The present review summarizes the proposed N-glycosylation pathway of S. acidocaldarius, describing the phenotypes of the mutants disrupted in N-glycan biosynthesis as well as giving insights into the archaeal O-linked and glycosylphosphatidylinositol anchor glycosylation process.

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Regulation of lamin properties and functions: does phosphorylation do it all?

Open Biology ◽

10.1098/rsob.150094 ◽

2015 ◽

Vol 5 (11) ◽

pp. 150094 ◽

Cited By ~ 32

Author(s):

Magdalena Machowska ◽

Katarzyna Piekarowicz ◽

Ryszard Rzepecki

Keyword(s):

Gene Expression ◽

Experimental Data ◽

Chromatin Structure ◽

In Silico ◽

Building Blocks ◽

Model Organisms ◽

Intracellular Signalling Pathway ◽

Evolutionarily Conserved ◽

Structure Regulation

The main functions of lamins are their mechanical and structural roles as major building blocks of the karyoskeleton. They are also involved in chromatin structure regulation, gene expression, intracellular signalling pathway modulation and development. All essential lamin functions seem to depend on their capacity for assembly or disassembly after the receipt of specific signals, and after specific, selective and precisely regulated interactions through their various domains. Reversible phosphorylation of lamins is crucial for their functions, so it is important to understand how lamin polymerization and interactions are modulated, and which sequences may undergo such modifications. This review combines experimental data with results of our in silico analyses focused on lamin phosphorylation in model organisms to show the presence of evolutionarily conserved sequences and to indicate specific in vivo phosphorylations that affect particular functions.

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Applying Cryo-X-ray Photoelectron Spectroscopy to Study the Surface Chemical Composition of Fungi and Viruses

Frontiers in Chemistry ◽

10.3389/fchem.2021.666853 ◽

2021 ◽

Vol 9 ◽

Author(s):

Andrey Shchukarev ◽

Emelie Backman ◽

Samuel Watts ◽

Stefan Salentinig ◽

Constantin F. Urban ◽

...

Keyword(s):

Chemical Composition ◽

Photoelectron Spectroscopy ◽

Biological Systems ◽

Building Blocks ◽

Model Organisms ◽

Surface Chemical ◽

Surface Layers ◽

Xps Spectra ◽

X Ray ◽

Surface Chemical Composition

Interaction between microorganisms and their surroundings are generally mediated via the cell wall or cell envelope. An understanding of the overall chemical composition of these surface layers may give clues on how these interactions occur and suggest mechanisms to manipulate them. This knowledge is key, for instance, in research aiming to reduce colonization of medical devices and device-related infections from different types of microorganisms. In this context, X-ray photoelectron spectroscopy (XPS) is a powerful technique as its analysis depth below 10 nm enables studies of the outermost surface structures of microorganism. Of specific interest for the study of biological systems is cryogenic XPS (cryo-XPS). This technique allows studies of intact fast-frozen hydrated samples without the need for pre-treatment procedures that may cause the cell structure to collapse or change due to the loss of water. Previously, cryo-XPS has been applied to study bacterial and algal surfaces with respect to their composition of lipids, polysaccharides and peptide (protein and/or peptidoglycan). This contribution focuses onto two other groups of microorganisms with widely different architecture and modes of life, namely fungi and viruses. It evaluates to what extent existing models for data treatment of XPS spectra can be applied to understand the chemical composition of their very different surface layers. XPS data from model organisms as well as reference substances representing specific building blocks of their surface were collected and are presented. These results aims to guide future analysis of the surface chemical composition of biological systems.

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Hybridization Capture Using RAD Probes (hyRAD), a New Tool for Performing Genomic Analyses on Collection Specimens

10.1101/025551 ◽

2015 ◽

Cited By ~ 1

Author(s):

Tomasz Suchan ◽

Camille Pitteloud ◽

Nadezhda S. Gerasimova ◽

Anna Kostikova ◽

Sarah Schmid ◽

...

Keyword(s):

Restriction Site ◽

Degraded Dna ◽

Model Organisms ◽

Butterfly Species ◽

Large Set ◽

Grasshopper Species ◽

Size Selection ◽

Restriction Sites ◽

Museum Samples ◽

Orthologous Loci

In the recent years, many protocols aimed at reproducibly sequencing reduced-genome subsets in non-model organisms have been published. Among them, RAD-sequencing is one of the most widely used. It relies on digesting DNA with specific restriction enzymes and performing size selection on the resulting fragments. Despite its acknowledged utility, this method is of limited use with degraded DNA samples, such as those isolated from museum specimens, as these samples are less likely to harbor fragments long enough to comprise two restriction sites making possible ligation of the adapter sequences (in the case of double-digest RAD) or performing size selection of the resulting fragments (in the case of single-digest RAD). Here, we address these limitations by presenting a novel method called hybridization RAD (hyRAD). In this approach, biotinylated RAD fragments, covering a random fraction of the genome, are used as baits for capturing homologous fragments from genomic shotgun sequencing libraries. This simple and cost-effective approach allows sequencing of orthologous loci even from highly degraded DNA samples, opening new avenues of research in the field of museum genomics. Not relying on the restriction site presence, it improves among-sample loci coverage. In a trial study, hyRAD allowed us to obtain a large set of orthologous loci from fresh and museum samples from a non-model butterfly species, with a high proportion of single nucleotide polymorphisms present in all eight analyzed specimens, including 58-year-old museum samples. The utility of the method was further validated using 49 museum and fresh samples of a Palearctic grasshopper species for which the spatial genetic structure was previously assessed using mtDNA amplicons. The application of the method is eventually discussed in a wider context. As it does not rely on the restriction site presence, it is therefore not sensitive to among-sample loci polymorphisms in the restriction sites that usually causes loci dropout. This should enable the application of hyRAD to analyses at broader evolutionary scales.

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Modular metabolite assembly in C. elegans depends on carboxylesterases and formation of lysosome-related organelles

10.1101/2020.08.22.262956 ◽

2020 ◽

Author(s):

Henry H. Le ◽

Chester J. J. Wrobel ◽

Sarah M. Cohen ◽

Jingfang Yu ◽

Heenam Park ◽

...

Keyword(s):

Life History ◽

Lipid Metabolism ◽

Amino Acid ◽

Building Blocks ◽

Signaling Molecules ◽

Selective Assembly ◽

C Elegans ◽

Comparative Metabolomics ◽

History Of ◽

Lysosome Related Organelles

AbstractSignaling molecules derived from attachment of diverse metabolic building blocks to ascarosides play a central role in the life history of C. elegans and other nematodes; however, many aspects of their biogenesis remain unclear. Using comparative metabolomics, we show that a pathway mediating formation of intestinal lysosome-related organelles (LROs) is required for biosynthesis of most modular ascarosides as well as previously undescribed modular glucosides. Similar to modular ascarosides, the modular glucosides are derived from highly selective assembly of moieties from nucleoside, amino acid, neurotransmitter, and lipid metabolism, suggesting that modular glucosides, like the ascarosides, may serve signaling functions. We further show that carboxylesterases that localize to intestinal organelles are required for the assembly of both modular ascarosides and glucosides via ester and amide linkages. Further exploration of LRO function and carboxylesterase homologs in C. elegans and other animals may reveal additional new compound families and signaling paradigms.Abstract Figure

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Cellular Effects of 2´,3´-Cyclic Nucleotide Monophosphates in Gram-Negative Bacteria

Journal of Bacteriology ◽

10.1128/jb.00208-21 ◽

2021 ◽

Author(s):

Yashasvika Duggal ◽

Jennifer E. Kurasz ◽

Benjamin M. Fontaine ◽

Nick J. Marotta ◽

Shikha S. Chauhan ◽

...

Keyword(s):

Cyclic Nucleotide ◽

Intracellular Signaling ◽

Biofilm Formation ◽

Second Messengers ◽

Rna Degradation ◽

Signaling Molecules ◽

Nucleotide Metabolism ◽

E Coli ◽

Cellular Processes ◽

Intracellular Signaling Molecules

Organismal adaptations to environmental stimuli are governed by intracellular signaling molecules such as nucleotide second messengers. Recent studies have identified functional roles for the non-canonical 2´,3´-cyclic nucleotide monophosphates (2´,3´-cNMPs) in both eukaryotes and prokaryotes. In Escherichia coli , 2´,3´-cNMPs are produced by RNase I-catalyzed RNA degradation, and these cyclic nucleotides modulate biofilm formation through unknown mechanisms. The present work dissects cellular processes in E. coli and Salmonella Typhimurium that are modulated by 2´,3´-cNMPs through the development of cell-permeable 2´,3´-cNMP analogs and a 2´,3´-cyclic nucleotide phosphodiesterase. Utilization of these chemical and enzymatic tools, in conjunction with phenotypic and transcriptomic investigations, identified pathways regulated by 2´,3´-cNMPs, including flagellar motility and biofilm formation, and by oligoribonucleotides with 3’-terminal 2´,3´-cyclic phosphates, including responses to cellular stress. Furthermore, interrogation of metabolomic and organismal databases has identified 2´,3´-cNMPs in numerous organisms and homologs of the E. coli metabolic proteins that are involved in key eukaryotic pathways. Thus, the present work provides key insights into the roles of these understudied facets of nucleotide metabolism and signaling in prokaryotic physiology and suggest broad roles for 2´,3´-cNMPs among bacteria and eukaryotes. IMPORTANCE Bacteria adapt to environmental challenges by producing intracellular signaling molecules which control downstream pathways and alter cellular processes for survival. Nucleotide second messengers serve to transduce extracellular signals and regulate a wide array of intracellular pathways. Recently, 2´,3´-cyclic nucleotide monophosphates (2´,3´-cNMPs) were identified for contributing to the regulation of cellular pathways in eukaryotes and prokaryotes. In this study we define previously unknown cell processes that are affected by fluctuating 2´,3´-cNMP levels or RNA oligomers with 2´,3´-cyclic phosphate termini in E. coli and Salmonella Typhimurium, providing a framework for studying novel signaling networks in prokaryotes. Furthermore, we utilize metabolomics databases to identify additional prokaryotic and eukaryotic species that generate 2´,3´-cNMPs as a resource for future studies.

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Single-molecule analysis reveals self assembly and nanoscale segregation of two distinct cavin subcomplexes on caveolae

eLife ◽

10.7554/elife.01434 ◽

2014 ◽

Vol 3 ◽

Cited By ~ 84

Author(s):

Yann Gambin ◽

Nicholas Ariotti ◽

Kerrie-Ann McMahon ◽

Michele Bastiani ◽

Emma Sierecki ◽

...

Keyword(s):

Single Molecule ◽

Self Assembly ◽

Mammalian Cells ◽

Protein Complexes ◽

Building Blocks ◽

Scaffolding Protein ◽

Cell Extracts ◽

Single Molecule Analysis ◽

Exquisite Specificity

In mammalian cells three closely related cavin proteins cooperate with the scaffolding protein caveolin to form membrane invaginations known as caveolae. Here we have developed a novel single-molecule fluorescence approach to directly observe interactions and stoichiometries in protein complexes from cell extracts and from in vitro synthesized components. We show that up to 50 cavins associate on a caveola. However, rather than forming a single coat complex containing the three cavin family members, single-molecule analysis reveals an exquisite specificity of interactions between cavin1, cavin2 and cavin3. Changes in membrane tension can flatten the caveolae, causing the release of the cavin coat and its disassembly into separate cavin1-cavin2 and cavin1-cavin3 subcomplexes. Each of these subcomplexes contain 9 ± 2 cavin molecules and appear to be the building blocks of the caveolar coat. High resolution immunoelectron microscopy suggests a remarkable nanoscale organization of these separate subcomplexes, forming individual striations on the surface of caveolae.

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