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Methanogenic archaea are main actors in the carbon cycle but are sensitive to reactive sulfite. Some methanogens use a sulfite detoxification system that combines an F 420 H 2 -oxidase with a sulfite reductase, both of which are proposed precursors of modern enzymes. Here, we present snapshots of this coupled system, named coenzyme F 420 -dependent sulfite reductase (Group I Fsr), obtained from two marine methanogens. Fsr organizes as a homotetramer, harboring an intertwined six-[4Fe–4S] cluster relay characterized by spectroscopy. The wire, spanning 5.4 nm, electronically connects the flavin to the siroheme center. Despite a structural architecture similar to dissimilatory sulfite reductases, Fsr shows a siroheme coordination and a reaction mechanism identical to assimilatory sulfite reductases. Accordingly, the reaction of Fsr is unidirectional, reducing sulfite or nitrite with F 420 H 2 . Our results provide structural insights into this unique fusion, in which a primitive sulfite reductase turns a poison into an elementary block of life. The F 420 -dependent sulfite reductase protects some methanogenic archaea by converting toxic sulfite. Structural analysis reveals how the two active centers are electro-connected and provides a plausible picture of a primitive sulfite reductase.

Methanothermococcus thermolithotrophicus is the only known methanogen that grows on sulfate as its sole sulfur source, uniquely uniting methanogenesis and sulfate reduction. Here we use physiological, biochemical and structural analyses to provide a snapshot of the complete sulfate reduction pathway of this methanogenic archaeon. We find that later steps in this pathway are catalysed by atypical enzymes. PAPS (3′-phosphoadenosine 5′-phosphosulfate) released by APS kinase is converted into sulfite and 3′-phosphoadenosine 5′-phosphate (PAP) by a PAPS reductase that is similar to the APS reductases of dissimilatory sulfate reduction. A non-canonical PAP phosphatase then hydrolyses PAP. Finally, the F 420 -dependent sulfite reductase converts sulfite to sulfide for cellular assimilation. While metagenomic and metatranscriptomic studies suggest that the sulfate reduction pathway is present in several methanogens, the sulfate assimilation pathway in M. thermolithotrophicus is distinct. We propose that this pathway was ‘mix-and-matched’ through the acquisition of assimilatory and dissimilatory enzymes from other microorganisms and then repurposed to fill a unique metabolic role. Structural and biochemical analyses show how one archaeon links both sulfur reduction and methane production.

Soil microorganisms collectively consume 70 million tonnes of atmospheric hydrogen (H 2 ) a year, regulating atmospheric composition and climate change. In turn, consuming this dependable trace gas enables these microorganisms to survive even when their preferred organic energy sources are exhausted. Despite the importance of H 2 consumption for soil biodiversity and atmospheric regulation, the signals and sensors that regulate this process remain to be understood. Here, we demonstrate that a model soil bacterium turns on the machinery required for atmospheric H 2 consumption in direct response to being limited by organic carbon availability, through the process of catabolite repression. Specifically, in the absence of a sensor of the organic carbon source glycerol, a H 2 -consuming hydrogenase is highly expressed and active. These findings suggest that organic carbon levels have a major role in regulating trace gas oxidation, with implications for predicting how trace gas consumption and soil biodiversity respond to environmental change.

Methanogenic archaea are main actors in the carbon cycle but are sensitive to reactive sulfite. Some methanogens use a sulfite detoxification system that combines an F H -oxidase with a sulfite reductase, both of which are proposed precursors of modern enzymes. Here, we present snapshots of this coupled system, named coenzyme F -dependent sulfite reductase (Group I Fsr), obtained from two marine methanogens. Fsr organizes as a homotetramer, harboring an intertwined six-[4Fe-4S] cluster relay characterized by spectroscopy. The wire, spanning 5.4 nm, electronically connects the flavin to the siroheme center. Despite a structural architecture similar to dissimilatory sulfite reductases, Fsr shows a siroheme coordination and a reaction mechanism identical to assimilatory sulfite reductases. Accordingly, the reaction of Fsr is unidirectional, reducing sulfite or nitrite with F H . Our results provide structural insights into this unique fusion, in which a primitive sulfite reductase turns a poison into an elementary block of life.

By growing on sulfate as the sole source of sulfur, Methanothermococcus thermolithotrophicus breaks a dogma: the ancient metabolic pathways methanogenesis and sulfate-reduction should not co-occur in one organism due to toxic intermediates and energetic barriers. Using a complementary approach of physiological, biochemical, and structural studies, we provide a snapshot of the complete sulfate-reduction pathway of the methanogenic archaeon. While the first two reactions proceed via an ATP-sulfurylase and APS-kinase, common to other organisms, the further steps are catalysed by non-canonical enzymes. 3'-phosphoadenosine-5'-phosphosulfate (PAPS) released by the APS-kinase is converted into sulfite and 3'-phosphoadenosine-5'-phosphate (PAP) by a new class of PAPS-reductase that shares high similarity with the APS-reductases involved in dissimilatory sulfate-reduction. The generated PAP is efficiently hydrolysed by a PAP-phosphatase that was likely derived from an RNA exonuclease. Finally, the F420-dependent sulfite-reductase converts sulfite to sulfide for cellular assimilation. While metagenomic and metatranscriptomic studies suggest that genes of the sulfate-reduction pathway are present in various methanogens, M. thermolithotrophicus uses a distinct way to assimilate sulfate. We propose that its entire sulfate-assimilation pathway was derived from a mix-and-match strategy in which the methanogen acquired assimilatory and dissimilatory enzymes from other microorganisms and shaped them to fit its physiological needs. Competing Interest Statement The authors have declared no competing interest.

The coenzyme F420-dependent sulfite reductase (Fsr group I) protects hydrogenotrophic methanogens, one of the main contributors in worldwide methane emission, from toxic sulfite. Fsr is a single peptide composed of a F420H2-oxidase and a novel class of sulfite reductase. Both catalytic domains have been proposed to be the ancestors of modern F420-oxido/reductases and dissimilatory/assimilatory sulfite reductases. Here, we describe the X-ray crystal structures of Fsr natively isolated from Methanocaldococcus jannaschii (MjFsr) and Methanothermococcus thermolithotrophicus (MtFsr), respectively refined to 2.30 Å and 1.55 Å resolution. In both organisms, Fsr oligomerizes as a 280-kDa homotetramer, where each siroheme–[4Fe–4S] is catalytically active, in contrast to dissimilatory homologues. The siroheme–[4Fe–4S], embedded in the sulfite reductase domain, is electronically connected to the flavin in the F420H2-oxidase domain by five [4Fe–4S]-clusters. EPR spectroscopy determined the redox potentials of these [4Fe–4S]2+/1+ clusters (−435 to -275 mV), through which electrons flow from FAD to the siroheme–[4Fe–4S]2+/1+ (siroheme, -114 mV; [4Fe–4S] -445 mV). The electron relay is mainly organized by two inserted ferredoxin modules, which stabilize the higher degree of oligomerization. While the F420H2-oxidase part is similar to the β-subunit of F420-reducing hydrogenases, the sulfite reductase domain is structurally analogous to dissimilatory sulfite reductases, whereas its siroheme–[4Fe–4S] cofactor is bound in the same way as in assimilatory ones. Accordingly, the reaction of MtFsr is unidirectional, reducing sulfite or nitrite with F420H2. Our results provide the first structural insights into this unique fusion, a snapshot of a primitive sulfite reductase that turns a poison into an elementary block of Life.

Consumption of atmospheric hydrogen (H2) enables diverse aerobic microorganisms to grow and persist in resource-deprived environments. In the aerobic saprophyte Mycobacterium smegmatis, hydrogen oxidation is catalyzed by two differentially expressed, high-affinity, oxygen-insensitive uptake hydrogenases, Huc and Hhy. Huc enables mixotrophic growth and facilitates the transition from growth to dormancy. Although the huc operon is known to be upregulated in response to organic carbon deprivation, the specific signals and regulators modulating its expression remain unresolved. Here, we show that GylR, a glycerol-3-phosphate-sensing regulator of glycerol metabolism, plays a role in repression of huc expression in response to the availability of glycerol but not other carbon sources. Based on proteomic analyses and activity assays, mutation or knockdown of gylR leads to enhanced Huc production and activity. GylR and other key catabolite repressor proteins (Crp1, Crp2) do not directly bind the huc operon, indicating repression is mediated by unidentified transcription factors, with GylR acting as an upstream sensor. Here, we present data that suggests atmospheric H2 oxidation is regulated in response to organic carbon source availability through the process of catabolite repression. By identifying a key signal that prompts atmospheric H2 oxidation, these findings advance understanding of how aerobic bacteria adapt to changing environmental conditions and suggest that organic carbon levels are a key factor regulating the main sink of atmospheric H2 in soils globally. Soil microorganisms collectively consume 70 million tonnes of atmospheric H2 a year, regulating atmospheric composition and climate change. In turn, consuming this dependable trace gas enables these microorganisms to survive even when their preferred organic energy sources are exhausted. Despite the importance of H2 consumption for soil biodiversity and atmospheric regulation, the signals and sensors that regulate this process remain to be understood. Here, we demonstrate that a model soil bacterium turns on the machinery required for atmospheric H2 consumption in direct response to being limited by organic carbon availability, through the process of catabolite repression. Specifically, in the absence of a sensor of the organic carbon source glycerol, a H2-consuming hydrogenase is highly expressed and active. These findings suggest that organic carbon levels have a major role in regulating trace gas oxidation, with implications for predicting how trace gas consumption and soil biodiversity respond to environmental change.

Terrestrial Antarctica harbors compositionally diverse and functionally distinct microbial life. Yet the ecological and evolutionary processes enabling these communities to adapt to the polyextreme conditions of the continent remain largely unknown. Here, we address how horizontal gene transfer (HGT) and de novo mutations influence adaptation of microbial communities in 16 proglacial and mountainous Antarctic soils, from a combination of short- and long-read datasets. Phylogenetic reconciliation and mobile genetic element analysis of 676 metagenome-assembled genomes show that HGT events occur frequently within these microbial communities. While the transferred genes are distributed across diverse functional categories, those involved in energy metabolism are exchanged at relatively higher frequency. The genes for aerotrophy, i.e. the consumption of atmospheric trace gases to provide energy, carbon, and hydration, are among the most frequently and widely disseminated. Approximately a quarter of all carbon monoxide (CO) dehydrogenases and [NiFe]-hydrogenases that catalyze atmospheric CO and hydrogen (H2) oxidation are predicted to be horizontally acquired and are often closely associated with mobile genetic elements. In parallel, analysis of polymorphisms in protein-encoding genes suggests widespread purifying selection, demonstrated by a predominance of synonymous mutations. This selection is particularly intense for aerotrophy genes, providing further evidence that this process is critical for microbial survival in Antarctica. The genetic variation of hydrogenases is tightly associated with their predicted protein structures, with intense selection acting on critical sites that preserve their stability and function in Antarctic environments. Together, these findings show that previously unrecognised eco-evolutionary dynamics shape the composition and function of Antarctic desert microbial communities, and confirm aerotrophy is a strongly selected and horizontally disseminated trait.

Rumen microbiota enable ruminants to grow on fibrous plant materials but also produce methane, driving 5% of global greenhouse gas emissions and leading to a loss of gross energy content. Methanogenesis inhibitors such as 3-nitrooxypropanol (3-NOP) decrease methane emissions in ruminants when supplemented in feed. Yet we lack a system-wide, species-resolved understanding of how the rumen microbiota remodels following inhibition and how this influences animal production. Here, we conducted a large-scale trial with 51 dairy calves to analyse microbiota responses to 3-NOP, pairing host performance, emissions, and nutritional profiles with genome-resolved metagenomic and metatranscriptomic data. 3-NOP supplementation decreased methane emissions by an average of 62%, modulated short-chain fatty acid and H2 levels, and did not affect dietary intake or animal performance. We created a rumen microbial genome catalogue with an unprecedented mapping rate. We observed a strong reduction of methanogens and stimulation of reductive acetogens, primarily novel uncultivated lineages such as Candidatus Faecousia. However, there was a shift in major fermentative communities away from acetate production in response to hydrogen gas accumulation. Thus, the divergent responses of the fermentative and hydrogenotrophic communities limit potential productivity gains from methane reduction. Reporting one of the largest reductions in methane emissions in a field trial to date, this study links ruminant greenhouse gas emissions and productivity to specific microbial species. These findings also emphasise the importance of microbiota-wide analysis for optimising methane mitigation strategies and identify promising strategies to simultaneously reduce emissions while increasing animal production. One strategy to increase the sustainability and productivity of livestock production is to modulate ruminant microbiota to produce absorbable nutrients rather than the potent greenhouse gas methane. Previous studies show supplementing feed with methanogenesis inhibitors such as 3-nitrooxypropanol reduces methane emissions, but also leads to inconsistent productivity gains. Here we report a definitive field trial, combining animal data, meta-omics, and structural modelling, to resolve the key microbes and pathways controlling nutrient and methane production in ruminants. We show that shifts in composition and gene expression of hydrogen-cycling microbes reduce emissions but limit productivity gains. These findings offer insights at unprecedented resolution, while the data and analytical framework provide valuable resources to develop solutions to enhance livestock productivity and sustainability.

Endothelial protein C receptor is shown to be the receptor for Plasmodium falciparum erythrocyte membrane protein 1 variants associated with severe malaria. Drug target in childhood malaria Severe childhood malaria, still causing about a million deaths every year, is triggered by the binding of red blood cells infected with the parasite Plasmodium falciparum to the walls of the host's blood vessels. P. falciparum erythrocyte membrane protein 1 (PfEMP1) containing domain cassettes 8 and 13 is known to be associated with severe malaria, and here Thomas Lavstsen and colleagues identify the receptor for PfEMP1 as endothelial protein C receptor (EPCR), a protein involved in regulating blood coagulation and the inflammatory response. This work could help to explain why some episodes of malaria are life-threatening and involve severe inflammation and suggests a target for future antimalarials. Sequestration of Plasmodium falciparum -infected erythrocytes in host blood vessels is a key triggering event in the pathogenesis of severe childhood malaria, which is responsible for about one million deaths every year 1 . Sequestration is mediated by specific interactions between members of the P. falciparum erythrocyte membrane protein 1 (PfEMP1) family and receptors on the endothelial lining 2 . Severe childhood malaria is associated with expression of specific PfEMP1 subtypes containing domain cassettes (DCs) 8 and 13 (ref. 3 ), but the endothelial receptor for parasites expressing these proteins was unknown 4 , 5 . Here we identify endothelial protein C receptor (EPCR), which mediates the cytoprotective effects of activated protein C 6 , as the endothelial receptor for DC8 and DC13 PfEMP1. We show that EPCR binding is mediated through the amino-terminal cysteine-rich interdomain region (CIDRα1) of DC8 and group A PfEMP1 subfamilies, and that CIDRα1 interferes with protein C binding to EPCR. This PfEMP1 adhesive property links P. falciparum cytoadhesion to a host receptor involved in anticoagulation and endothelial cytoprotective pathways, and has implications for understanding malaria pathology and the development of new malaria interventions.