Several of the metabolic enzymes in methanotrophic bacteria rely on metals for both their expression and their catalysis. The MxaFI methanol dehydrogenase enzyme complex uses calcium as a cofactor to ...oxidize methanol, while the alternative methanol dehydrogenase XoxF uses lanthanide metals such as lanthanum and cerium for the same function. Lanthanide metals, abundant in the earth's crust, strongly repress the transcription of
yet activate the transcription of
This regulatory program, called the "lanthanide switch," is central to methylotrophic metabolism, but only some of its components are known. To uncover additional components of the lanthanide switch, we developed a chemical mutagenesis system in the type I gammaproteobacterial methanotroph "
" 5GB1C and designed a selection system for mutants unable to repress the
promoter in the presence of lanthanum. Whole-genome resequencing for multiple lanthanide switch mutants identified several unique point mutations in a single gene encoding a TonB-dependent receptor, which we have named LanA. The LanA TonB-dependent receptor is absolutely required for the lanthanide switch and controls the expression of a small set of genes. While mutation of the
gene does not affect the amount of cell-associated lanthanum, it is essential for growth in the absence of the MxaF methanol dehydrogenase, suggesting that LanA is involved in lanthanum uptake to supply the XoxF methanol dehydrogenase with its critical metal ion cofactor. The discovery of this novel component of the lanthanide regulatory system highlights the complexity of this circuit and suggests that further components are likely involved.
Lanthanide metals, or rare earth elements, are abundant in nature and used heavily in technological devices. Biological interactions with lanthanides are just beginning to be unraveled. Until very recently, microbial mechanisms of lanthanide metal interaction and uptake were unknown. The TonB-dependent receptor LanA is the first lanthanum receptor identified in a methanotroph. Sequence homology searches with known metal transporters and regulators could not be used to identify LanA or other lanthanide metal switch components, and this method for mutagenesis and selection was required to identify the receptor. This work advances the knowledge of microbe-metal interactions in environmental niches that impact atmospheric methane levels and are thus relevant to climate change.
5GB1C, a fast-growing gammaproteobacterial methanotroph, is equipped with two glycolytic pathways, the Entner-Doudoroff (ED) pathway and the Embden-Meyerhof-Parnas (EMP) pathway. Metabolic flux ...analysis and
C-labeling experiments have shown the EMP pathway is the principal glycolytic route in
5GB1C, while the ED pathway appears to be metabolically and energetically insignificant. However, it has not been possible to obtain a null mutant in the
-
genes encoding the two unique enzymatic reactions in the ED pathway, suggesting the ED pathway may be essential for
5GB1C growth. In this study, the inducible P
promoter was used to manipulate gene expression of
, and in addition, the expression of these two genes was separated from that of a downstream
gene. The resulting strain shows arabinose-dependent growth that correlates with ED pathway activity, with normal growth achieved in the presence of ∼0.1 g/liter arabinose. Flux balance analysis shows that
5GB1C with a strong ED pathway has a reduced energy budget, thereby limiting the mutant growth at a high concentration of arabinose. Collectively, our study demonstrates that the ED pathway is essential for
5GB1C. However, no known mechanism can directly explain the essentiality of the ED pathway, and thus, it may have a yet unknown regulatory role required for sustaining a healthy and functional metabolism in this bacterium.
The gammaproteobacterial methanotrophs possess a unique central metabolic architecture where methane and other reduced C
carbon sources are assimilated through the ribulose monophosphate cycle. Although efforts have been made to better understand methanotrophic metabolism in these bacteria via experimental and computational approaches, many questions remain unanswered. One of these is the essentiality of the ED pathway for
5GB1C, as current results appear contradictory. By creating a construct with
and
genes controlled by P
and P
, respectively, we demonstrated the essentiality of the ED pathway for this obligate methanotroph. It is also demonstrated that these genetic tools are applicable to
5GB1C and that expression of the target genes can be tightly controlled across an extensive range. Our study adds to the expanding knowledge of methanotrophic metabolism and practical approaches to genetic manipulation for obligate methanotrophs.
The rapid increase of the potent greenhouse gas methane in the atmosphere creates great urgency to develop and deploy technologies for methane mitigation. One approach to removing methane is to use ...bacteria for which methane is their carbon and energy source (methanotrophs). Such bacteria naturally convert methane to CO
2
and biomass, a value-added product and a cobenefit of methane removal. Typically, methanotrophs grow best at around 5,000 to 10,000 ppm methane, but methane in the atmosphere is 1.9 ppm. Air above emission sites such as landfills, anaerobic digestor effluents, rice paddy effluents, and oil and gas wells contains elevated methane in the 500 ppm range. If such sites are targeted for methane removal, technology harnessing aerobic methanotroph metabolism has the potential to become economically and environmentally viable. The first step in developing such methane removal technology is to identify methanotrophs with enhanced ability to grow and consume methane at 500 ppm and lower. We report here that some existing methanotrophic strains grow well at 500 ppm methane, and one of them,
Methylotuvimicrobium buryatense
5GB1C, consumes such low methane at enhanced rates compared to previously published values. Analyses of bioreactor-based performance and RNAseq-based transcriptomics suggest that this ability to utilize low methane is based at least in part on extremely low non-growth-associated maintenance energy and on high methane specific affinity. This bacterium is a candidate to develop technology for methane removal at emission sites. If appropriately scaled, such technology has the potential to slow global warming by 2050.
Engineering microorganisms into biological factories that convert renewable feedstocks into valuable materials is a major goal of synthetic biology; however, for many nonmodel organisms, we do not ...yet have the genetic tools, such as suites of strong promoters, necessary to effectively engineer them. In this work, we developed a computational framework that can leverage standard RNA-seq data sets to identify sets of constitutive, strongly expressed genes and predict strong promoter signals within their upstream regions. The framework was applied to a diverse collection of RNA-seq data measured for the methanotroph Methylotuvimicrobium buryatense 5GB1 and identified 25 genes that were constitutively, strongly expressed across 12 experimental conditions. For each gene, the framework predicted short (27–30 nucleotide) sequences as candidate promoters and derived −35 and −10 consensus promoter motifs (TTGACA and TATAAT, respectively) for strong expression in M. buryatense. This consensus closely matches the canonical E. coli sigma-70 motif and was found to be enriched in promoter regions of the genome. A subset of promoter predictions was experimentally validated in a XylE reporter assay, including the consensus promoter, which showed high expression. The pmoC, pqqA, and ssrA promoter predictions were additionally screened in an experiment that scrambled the −35 and −10 signal sequences, confirming that transcription initiation was disrupted when these specific regions of the predicted sequence were altered. These results indicate that the computational framework can make biologically meaningful promoter predictions and identify key pieces of regulatory systems that can serve as foundational tools for engineering diverse microorganisms for biomolecule production.
Full text
Available for:
IJS, KILJ, NUK, PNG, UL, UM
In a study involving 3975 health care personnel, first responders, and other essential workers, the effectiveness of mRNA vaccines against SARS-CoV-2 infection was 91% with full vaccination. The ...vaccines attenuated the viral RNA load, febrile symptoms, and illness duration among those who became infected despite vaccination.
Group 2 innate lymphoid cells (ILC2s) are essential to maintain tissue homeostasis. In cancer, ILC2s can harbor both pro-tumorigenic and anti-tumorigenic functions, but we know little about their ...underlying mechanisms or whether they could be clinically relevant or targeted to improve patient outcomes. Here, we found that high ILC2 infiltration in human melanoma was associated with a good clinical prognosis. ILC2s are critical producers of the cytokine granulocyte-macrophage colony-stimulating factor, which coordinates the recruitment and activation of eosinophils to enhance antitumor responses. Tumor-infiltrating ILC2s expressed programmed cell death protein-1, which limited their intratumoral accumulation, proliferation and antitumor effector functions. This inhibition could be overcome in vivo by combining interleukin-33-driven ILC2 activation with programmed cell death protein-1 blockade to significantly increase antitumor responses. Together, our results identified ILC2s as a critical immune cell type involved in melanoma immunity and revealed a potential synergistic approach to harness ILC2 function for antitumor immunotherapies.
Full text
Available for:
GEOZS, IJS, IMTLJ, KISLJ, NLZOH, NUK, OILJ, PNG, SAZU, SBCE, SBMB, UL, UM, UPUK, ZAGLJ
Should governments use a discount rate that declines over time when evaluating the future
benefits and costs of public projects? The argument for using a declining discount rate
(DDR) is simple: if ...the discount rates that will be applied in the future are uncertain
but positively correlated, and if the analyst can assign probabilities to these discount
rates, then the result will be a declining schedule of certainty-equivalent discount
rates. There is a growing empirical literature that estimates models of long-term interest
rates and uses them to forecast the DDR schedule. However, this literature has been
criticized because it lacks a connection to the theory of project evaluation. In
benefit-cost analysis, the net benefits of a project in year t (in
consumption units) are discounted to the present at the rate at which society would trade
consumption in year t for consumption in the present. With simplifying
assumptions, this leads to the Ramsey discounting formula, which results in a declining
certainty-equivalent discount rate if the rate of growth in consumption is uncertain and
if shocks to consumption are correlated over time. We conclude that the arguments in favor
of a DDR are compelling and thus merit serious consideration by regulatory agencies in the
United States. (JEL: D61)
Epigenetic markings acquired in early life may have phenotypic consequences later in development through their role in transcriptional regulation with relevance to the developmental origins of ...diseases including obesity. The goal of this study was to investigate whether DNA methylation levels at birth are associated with body size later in childhood.
A study design involving two birth cohorts was used to conduct transcription profiling followed by DNA methylation analysis in peripheral blood. Gene expression analysis was undertaken in 24 individuals whose biological samples and clinical data were collected at a mean ± standard deviation (SD) age of 12.35 (0.95) years, the upper and lower tertiles of body mass index (BMI) were compared with a mean (SD) BMI difference of 9.86 (2.37) kg/m(2). This generated a panel of differentially expressed genes for DNA methylation analysis which was then undertaken in cord blood DNA in 178 individuals with body composition data prospectively collected at a mean (SD) age of 9.83 (0.23) years. Twenty-nine differentially expressed genes (>1.2-fold and p<10(-4)) were analysed to determine DNA methylation levels at 1-3 sites per gene. Five genes were unmethylated and DNA methylation in the remaining 24 genes was analysed using linear regression with bootstrapping. Methylation in 9 of the 24 (37.5%) genes studied was associated with at least one index of body composition (BMI, fat mass, lean mass, height) at age 9 years, although only one of these associations remained after correction for multiple testing (ALPL with height, p(Corrected) = 0.017).
DNA methylation patterns in cord blood show some association with altered gene expression, body size and composition in childhood. The observed relationship is correlative and despite suggestion of a mechanistic epigenetic link between in utero life and later phenotype, further investigation is required to establish causality.
Full text
Available for:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Stars with zero-age main sequence masses between 140 and 260 M ⊙ are thought to explode as pair-instability supernovae (PISNe). During their thermonuclear runaway, PISNe can produce up to several ...tens of solar masses of radioactive nickel, resulting in luminous transients similar to some superluminous supernovae (SLSNe). Yet, no unambiguous PISN has been discovered so far. SN 2018ibb is a hydrogen-poor SLSN at z = 0.166 that evolves extremely slowly compared to the hundreds of known SLSNe. Between mid 2018 and early 2022, we monitored its photometric and spectroscopic evolution from the UV to the near-infrared (NIR) with 2–10 m class telescopes. SN 2018ibb radiated > 3 × 10 51 erg during its evolution, and its bolometric light curve reached > 2 × 10 44 erg s −1 at its peak. The long-lasting rise of > 93 rest-frame days implies a long diffusion time, which requires a very high total ejected mass. The PISN mechanism naturally provides both the energy source ( 56 Ni) and the long diffusion time. Theoretical models of PISNe make clear predictions as to their photometric and spectroscopic properties. SN 2018ibb complies with most tests on the light curves, nebular spectra and host galaxy, and potentially all tests with the interpretation we propose. Both the light curve and the spectra require 25–44 M ⊙ of freshly nucleosynthesised 56 Ni, pointing to the explosion of a metal-poor star with a helium core mass of 120–130 M ⊙ at the time of death. This interpretation is also supported by the tentative detection of Co II λ 1.025 μm, which has never been observed in any other PISN candidate or SLSN before. We observe a significant excess in the blue part of the optical spectrum during the nebular phase, which is in tension with predictions of existing PISN models. However, we have compelling observational evidence for an eruptive mass-loss episode of the progenitor of SN 2018ibb shortly before the explosion, and our dataset reveals that the interaction of the SN ejecta with this oxygen-rich circumstellar material contributed to the observed emission. That may explain this specific discrepancy with PISN models. Powering by a central engine, such as a magnetar or a black hole, can be excluded with high confidence. This makes SN 2018ibb by far the best candidate for being a PISN, to date.
Full text
Available for:
FMFMET, NUK, UL, UM, UPUK
Mutations in the skeletal muscle Ca
release channel, the type 1 ryanodine receptor (RYR1), cause malignant hyperthermia susceptibility (MHS) and a life-threatening sensitivity to heat, which is most ...severe in children. Mice with an MHS-associated mutation in Ryr1 (Y524S, YS) display lethal muscle contractures in response to heat. Here we show that the heat response in the YS mice is exacerbated by brown fat adaptive thermogenesis. In addition, the YS mice have more brown adipose tissue thermogenic capacity than their littermate controls. Blood lactate levels are elevated in both heat-sensitive MHS patients with RYR1 mutations and YS mice due to Ca
driven increases in muscle metabolism. Lactate increases brown adipogenesis in both mouse and human brown preadipocytes. This study suggests that simple lifestyle modifications such as avoiding extreme temperatures and maintaining thermoneutrality could decrease the risk of life-threatening responses to heat and exercise in individuals with RYR1 pathogenic variants.
PDF
