Effects of H ₂ and Formate on Growth Yield and Regulation of Methanogenesis in Methanococcus maripaludis

Abstract

bHydrogenotrophic methanogenic Archaea are defined by an H 2 requirement for growth. Despite this requirement, many hydrogenotrophs are also capable of growth with formate as an electron donor for methanogenesis. While certain responses of these organisms to hydrogen availability have been characterized, responses to formate starvation have not been reported. Here we report that during continuous culture of Methanococcus maripaludis under defined nutrient conditions, growth yields relative to methane production decreased markedly with either H 2 excess or formate excess. Analysis of the growth yields of several mutants suggests that this phenomenon occurs independently of the storage of intracellular carbon or a transcriptional response to methanogenesis. Using microarray analysis, we found that the expression of genes encoding coenzyme F 420 -dependent steps of methanogenesis, including one of two formate dehydrogenases, increased with H 2 starvation but with formate occurred at high levels regardless of limitation or excess. One gene, encoding H 2 -dependent methylene-tetrahydromethanopterin dehydrogenase, decreased in expression with either H 2 limitation or formate limitation. Expression of genes for the second formate dehydrogenase, molybdenum-dependent formylmethanofuran dehydrogenase, and molybdenum transport increased specifically with formate limitation. Of the two formate dehydrogenases, only the first could support growth on formate in batch culture where formate was in excess.

Carbohydrate Metabolism in Archaea: Current Insights into Unusual Enzymes and Pathways and Their Regulation

“…In contrast, a recent genetic/transcriptomic study by Costa et al (578) rather supports the idea that energy spilling (i.e., futile cycling) might play an important physiological role in Mco. maripaludis under conditions of H 2 and formate excess.…”

A systems level predictive model for global gene regulation of methanogenesis in a hydrogenotrophic methanogen

“…In contrast, CH 4 production increased and decreased much more rapidly. This response reflects a marked difference in growth yield on a per-CH 4 basis under the two H 2 conditions (Costa et al 2013b). 2.…”

“…These experiments included 52 transcriptome profiles from gene deletion strains (Fig. 1C) and four from steadystate cultures grown with formate as an electron donor ''without'' H 2 (Costa et al 2013b). Culture conditions and samples for transcriptome analysis are summarized in Supplemental Table S4.…”

“…The fragments were digested with AscI, ligated, PCR amplified, cloned into pCRuptneo, and introduced into MM901 as above. To make MM901DMMP1447 PnifTeha (strain MM1322) (Costa et al 2013b), regions flanking MMP1447 were PCR amplified and cloned into the plasmid pHW40 containing Pnif (Chaban et al 2007). The product was PCR amplified, ligated into pCRuptneo, and introduced into MM901 as above.…”

“…Along with eight transcriptome data sets from our previous study (Yoon et al 2011), a total of 58 transcriptome profiles of wild-type M. maripaludis MM901 were analyzed using the cMonkey biclustering algorithm ) and the Inferelator algorithm , in order to infer the set of environmental factors (EFs) and transcriptional factors (TFs, sequence-specific DNA-binding proteins that mediate regulation of mRNA transcription) that accurately modeled the dynamic transcriptional changes of genes within each bicluster. The EGRIN model was validated using 56 transcriptome data sets that were not used for the model construction: 52 new transcriptome data sets from several genetic backgrounds that modulated methanogenesis in well-understood schemes and four data sets from our previous work (Costa et al 2013b). We demonstrated that this EGRIN model accurately predicted transcriptional responses of all genes in the network and can be used as a framework to formulate novel hypotheses regarding gene functions and regulation.…”