Model Organisms To Study Methanogenesis, a Uniquely Archaeal Metabolism

Costa, Kyle C.; Whitman, William B.

doi:10.1128/jb.00115-23

Cited by 11 publications

(4 citation statements)

References 188 publications

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“…In these cases, and in other instances where the research goal is not highly specific for one particular methanogen, the selection of a proper methanogenic host is the first step in the genetic engineering process. A recent review by Costa and Whitman ( 32 ) provides a comprehensive overview of current model methanogen species. The present review instead aims to focus on describing current genetic tools for methanogens.…”

Section: Host Methanogensmentioning

confidence: 99%

Teaching old dogs new tricks: genetic engineering methanogens

Myers,

Dykstra

2024

Appl Environ Microbiol

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Methanogenic archaea, which are integral to global carbon and nitrogen cycling, currently face challenges in genetic manipulation due to unique physiology and limited genetic tools. This review provides a survey of current and past developments in the genetic engineering of methanogens, including selection and counterselection markers, reporter systems, shuttle vectors, mutagenesis methods, markerless genetic exchange, and gene expression control. This review discusses genetic tools and emphasizes challenges tied to tool scarcity for specific methanogenic species. Mutagenesis techniques for methanogens, including physicochemical, transposon-mediated, liposome-mediated mutagenesis, and natural transformation, are outlined, along with achievements and challenges. Markerless genetic exchange strategies, such as homologous recombination and CRISPR/Cas-mediated genome editing, are also detailed. Finally, the review concludes by examining the control of gene expression in methanogens. The information presented underscores the urgent need for refined genetic tools in archaeal research. Despite historical challenges, recent advancements, notably CRISPR-based systems, hold promise for overcoming obstacles, with implications for global health, agriculture, climate change, and environmental engineering. This comprehensive review aims to bridge existing gaps in the literature, guiding future research in the expanding field of archaeal genetic engineering.

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Section: Host Methanogensmentioning

confidence: 99%

Teaching old dogs new tricks: genetic engineering methanogens

Myers,

Dykstra

2024

Appl Environ Microbiol

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“…The study of model organisms is key to overcoming the challenges associated with characterizing archaeal physiology. Methanococcus maripaludis is a model (2) with well-established genetic tools for understanding archaeal biology, methanogenesis, and diazotrophy. However, even in this well-studied organism, ~35% of proteins have RefSeq annotations of either "hypothetical protein", "membrane protein", or belonging to a family rather than having specific functional assignments.…”

Section: Introductionmentioning

confidence: 99%

High-throughput genetics enables identification of nutrient utilization and accessory energy metabolism genes in a model methanogen

Day,

Carlson,

Fonseca

et al. 2024

Preprint

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Archaea are widespread in the environment and play fundamental roles in diverse ecosystems; however, characterization of their unique biology requires advanced tools. This is particularly challenging when characterizing gene function. Here, we generate randomly-barcoded transposon libraries in the model methanogenic archaeon Methanococcus maripaludis and use high-throughput growth methods to conduct fitness assays (RB-TnSeq) across over 100 unique growth conditions. Using our approach, we identified new genes involved in nutrient utilization and response to oxidative stress. We identified novel genes for the usage of diverse nitrogen sources in M. maripaludis including a putative regulator of alanine deamination and molybdate transporters important for nitrogen fixation. Furthermore, leveraging the fitness data, we inferred that M. maripaludis can utilize additional nitrogen sources including L-glutamine, D-glucuronamide, and adenosine. Under autotrophic growth conditions, we identified a gene encoding a domain of unknown function (DUF166) that is important for fitness and hypothesize it has an accessory role in carbon dioxide assimilation. Finally, comparing fitness costs of oxygen versus sulfite stress, we identified a previously uncharacterized class of dissimilatory sulfite reductase-like proteins (Dsr-LP, group IIId) that is important during growth in the presence of sulfite. When overexpressed, Dsr-LP conferred sulfite resistance and enabled use of sulfite as the sole sulfur source. The high-throughput approach employed here allowed for generation of a large-scale dataset that can be used as a resource to further understand gene function and metabolism in the archaeal domain.

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“…Methanogenic archaea are single-celled organisms, which were first discovered about a century ago ( 1 , 2 ). Recently, methanogens have received increased research interest because of a key role in climate change via methane emissions, which represent a greenhouse gas ( 3 ).…”

Section: Introductionmentioning

confidence: 99%

The novel regulator HdrR controls the transcription of the heterodisulfide reductase operon hdrBCA in Methanosarcina barkeri

Zhang,

Chen,

Wang

et al. 2024

Appl Environ Microbiol

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Methanogenic archaea play a key role in the global carbon cycle because these microorganisms remineralize organic compounds in various anaerobic environments. The microorganism Methanosarcina barkeri is a metabolically versatile methanogen, which can utilize acetate, methanol, and H 2 /CO 2 to synthesize methane. However, the regulatory mechanisms underlying methanogenesis for different substrates remain unknown. In this study, RNA-seq analysis was used to investigate M. barkeri growth and gene transcription under different substrate regimes. According to the results, M. barkeri showed the best growth under methanol, followed by H 2 /CO 2 and acetate, and these findings corresponded well with the observed variations in genes transcription abundance for different substrates. In addition, we identified a novel regulator, MSBRM_RS03855 (designated as HdrR), which specifically activates the transcription of the heterodisulfide reductase hdrBCA operon in M. barkeri . HdrR was able to bind to the hdrBCA operon promoter to regulate transcription. Furthermore, the structural model analyses revealed a helix-turn-helix domain, which is likely involved in DNA binding. Taken together, HdrR serves as a model to reveal how certain regulatory factors control the expression of key enzymes in the methanogenic pathway. IMPORTANCE The microorganism Methanosarcina barkeri has a pivotal role in the global carbon cycle and contributes to global temperature homeostasis. The consequences of biological methanogenesis are far-reaching, including impacts on atmospheric methane and CO 2 concentrations, agriculture, energy production, waste treatment, and human health. As such, reducing methane emissions is crucial to meeting set climate goals. The methanogenic activity of certain microorganisms can be drastically reduced by inhibiting the transcription of the hdrBCA operon, which encodes heterodisulfide reductases. Here, we provide novel insight into the mechanisms regulating hdrBCA operon transcription in the model methanogen M. barkeri . The results clarified that HdrR serves as a regulator of heterodisulfide reductase hdrBCA operon transcription during methanogenesis, which expands our understanding of the unique regulatory mechanisms that govern methanogenesis. The findings presented in this study can further our understanding of how genetic regulation can effectively reduce the methane emissions caused by methanogens.

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Model Organisms To Study Methanogenesis, a Uniquely Archaeal Metabolism

Cited by 11 publications

References 188 publications

Teaching old dogs new tricks: genetic engineering methanogens

Teaching old dogs new tricks: genetic engineering methanogens

High-throughput genetics enables identification of nutrient utilization and accessory energy metabolism genes in a model methanogen

The novel regulator HdrR controls the transcription of the heterodisulfide reductase operon hdrBCA in Methanosarcina barkeri

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