skip to main content
US FlagAn official website of the United States government
dot gov icon
Official websites use .gov
A .gov website belongs to an official government organization in the United States.
https lock icon
Secure .gov websites use HTTPS
A lock ( lock ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.


Title: Alternative carbon sources for the production of plant cellular agriculture: a case study on acetate
Plant cellular agriculture aims to disrupt the way plant derived products are produced. Plant cell cultures are typically grown with sucrose as the primary carbon and energy source, but alternative carbon sources may have advantages over sucrose including less strain on food systems, lower costs, and more sustainable sourcing. Here we review carbon and energy sources that may serve as alternatives to sucrose in the cultivation of plant cell cultures. We identified acetate as a promising candidate and took the first steps to evaluate its potential for use in growing tobacco plant cell cultures. When added to media containing sucrose, acetate concentrations above 8 mM completely inhibit growth. Lower concentrations of acetate (2-4 mM) can support an increase in dry weight without sucrose but do not provide enough energy for substantial growth.13C labeling indicates that tobacco plant cell cultures can incorporate carbon from exogenous acetate into proteins and carbohydrates. Analysis of transcriptome data showed that genes encoding glyoxylate cycle enzymes are expressed at very low levels compared to genes from the TCA cycle and glycolysis. Adaptive laboratory evolution experiments were able to increase tobacco cell cultures tolerance to acetate, demonstrating the potential for this type of approach going forward. Overall, our results indicate that acetate can be metabolized by plant cell cultures and suggest that further adaptive laboratory evolution or strain engineering efforts may enable acetate to serve as a sole carbon and energy source for tobacco plant cell cultures. This assessment of acetate provides a framework for evaluating other carbon and energy sources for plant cell cultures, efforts that will help reduce the costs and environmental impact, and increase the commercial potential of plant cellular agriculture.  more » « less
Award ID(s):
1922642
PAR ID:
10635682
Author(s) / Creator(s):
; ; ; ;
Publisher / Repository:
Frontiers Plant Sci
Date Published:
Journal Name:
Frontiers in Plant Science
Volume:
14
ISSN:
1664-462X
Format(s):
Medium: X
Sponsoring Org:
National Science Foundation
More Like this
  1. Abstract We report the isolation a halophilic bacterium that degrades both aromatic and aliphatic hydrocarbons as the sole sources of carbon at high salinity from produced water. Phylogenetic analysis of 16S rRNA-gene sequences shows the isolate is a close relative of Modicisalibacter tunisiensis isolated from an oil-field water in Tunisia. We designate our isolate as Modicisalibacter sp. strain Wilcox. Genome analysis of strain Wilcox revealed the presence of a repertoire of genes involved in the metabolism of aliphatic and aromatic hydrocarbons. Laboratory culture studies corroborated the predicted hydrocarbon degradation potential. The strain degraded benzene, toluene, ethylbenzene, and xylenes at salinities ranging from 0.016 to 4.0 M NaCl, with optimal degradation at 1 M NaCl. Also, the strain degraded phenol, benzoate, biphenyl and phenylacetate as the sole sources of carbon at 2.5 M NaCl. Among aliphatic compounds, the strain degraded n-decane and n-hexadecane as the sole sources of carbon at 2.5 M NaCl. Genome analysis also predicted the presence of many heavy metal resistance genes including genes for metal efflux pumps, transport proteins, and enzymatic detoxification. Overall, due to its ability to degrade many hydrocarbons and withstand high salt and heavy metals, strain Wilcox may prove useful for remediation of produced waters. 
    more » « less
  2. null (Ed.)
    Sucrose is among the main products of photosynthesis that are deemed necessary for plant growth and survival. It is produced in the mesophyll cells of leaves and translocated to different parts of the plant through the phloem. Progress in understanding this transport process remains fraught with experimental difficulties, thereby prompting interest in theoretical approaches and laboratory studies. The Münch pressure and mass flow model is one of the accepted hypotheses describing the physics of sucrose transport in the phloem. It is based on osmosis creating an energy potential difference between the source and the sink. The flow responding to this energy potential is assumed laminar and described by the Hagen–Poiseuille equation. This study revisits such osmotically driven flows in tubes with membrane walls by including the effects of Taylor dispersion on mass transport. This effect has been overlooked in phloem flow studies. Taylor dispersion can increase the effective transport of solutes by increasing the apparent diffusion coefficient. It is shown that, in addition to the conventional diffusive correction derived for impermeable tube walls, a new adjustment to the mean advective terms arises because of osmotic effects. Because the molecular Schmidt number is very large for sucrose in water, the sucrose front speed and travel times have a direct dependence on the Péclet number for different ranges of the Münch number. This study establishes upper limits on expected Taylor dispersion enhancement of sucrose transport. 
    more » « less
  3. Abstract Evolutionary adaptation increases the fitness of a species in its environment. It can occur through rewiring of gene regulatory networks, such that an organism responds appropriately to environmental changes. We investigated whether sirtuin deacetylases, which repress transcription and require NAD+ for activity, serve as transcriptional rewiring points that facilitate the evolution of potentially adaptive traits. If so, bringing genes under the control of sirtuins could enable organisms to mount appropriate responses to stresses that decrease NAD+ levels. To explore how the genomic targets of sirtuins shift over evolutionary time, we compared two yeast species, Saccharomyces cerevisiae and Kluyveromyces lactis, that display differences in cellular metabolism and life cycle timing in response to nutrient availability. We identified sirtuin-regulated genes through a combination of chromatin immunoprecipitation and RNA expression. In both species, regulated genes were associated with NAD+ homeostasis, mating, and sporulation, but the specific genes differed. In addition, regulated genes in K. lactis were associated with other processes, including utilization of nonglucose carbon sources, detoxification of arsenic, and production of the siderophore pulcherrimin. Consistent with the species-restricted regulation of these genes, sirtuin deletion affected relevant phenotypes in K. lactis but not S. cerevisiae. Finally, sirtuin-regulated gene sets were depleted for broadly conserved genes, consistent with sirtuins regulating processes restricted to a few species. Taken together, these results are consistent with the notion that sirtuins serve as rewiring points that allow species to evolve distinct responses to low NAD+ stress. 
    more » « less
  4. Tringe, Susannah Green (Ed.)
    ABSTRACT Methanotrophic bacteria play a vital role in the biogeochemical carbon cycle due to their unique ability to use CH4as a carbon and energy source. Evidence suggests that some methanotrophs, includingMethylococcus capsulatus, can also use CO2as a carbon source, making these bacteria promising candidates for developing biotechnologies targeting greenhouse gas capture and mitigation. However, a deeper understanding of the dual CH4and CO2metabolism is needed to guide methanotroph strain improvements and realize their industrial utility. In this study, we show thatM. capsulatusexpresses five carbonic anhydrase (CA) isoforms, one α-CA, one γ-CA, and three β-CAs, that play a role in its inorganic carbon metabolism and CO2-dependent growth. The CA isoforms are differentially expressed, and transcription of all isoform genes is induced in response to CO2limitation. CA null mutant strains exhibited markedly impaired growth compared to an isogenic wild-type control, suggesting that the CA isoforms have independent, non-redundant roles inM. capsulatusmetabolism and physiology. Overexpression of some, but not all, CA isoforms improved bacterial growth kinetics and decreased CO2evolution from CH4-consuming cultures. Notably, we developed an engineered methanotrophic biocatalyst overexpressing the native α-CA and β-CA with a 2.5-fold improvement in the conversion of CH4to biomass. Given that product yield is a significant cost driver of methanotroph-based bioprocesses, the engineered strain developed here could improve the economics of CH4biocatalysis, including the production of single-cell protein from natural gas or anaerobic digestion-derived biogas.IMPORTANCEMethanotrophs transform CH4into CO2and multi-carbon compounds, so they play a critical role in the global carbon cycle and are of interest for biotechnology applications. Some methanotrophs, includingMethylococcus capsulatus, can also use CO2as a carbon source, but this dual one-carbon metabolism is incompletely understood. In this study, we show thatM. capsulatuscarbonic anhydrases are critical for this bacterium to optimally utilize CO2. We developed an engineered strain with improved CO2utilization capacity that increased the overall carbon conversion to cell biomass. The improvements to methanotroph-based product yields observed here are expected to reduce costs associated with CH4conversion bioprocesses. 
    more » « less
  5. ABSTRACT Although alcohols are toxic to many microorganisms, they are good carbon and energy sources for some bacteria, including many pseudomonads. However, most studies that have examined chemosensory responses to alcohols have reported that alcohols are sensed as repellents, which is consistent with their toxic properties. In this study, we examined the chemotaxis of Pseudomonas putida strain F1 to n -alcohols with chain lengths of 1 to 12 carbons. P. putida F1 was attracted to all n -alcohols that served as growth substrates (C 2 to C 12 ) for the strain, and the responses were induced when cells were grown in the presence of alcohols. By assaying mutant strains lacking single or multiple methyl-accepting chemotaxis proteins, the receptor mediating the response to C 2 to C 12 alcohols was identified as McfP, the ortholog of the P. putida strain KT2440 receptor for C 2 and C 3 carboxylic acids. Besides being a requirement for the response to n -alcohols, McfP was required for the response of P. putida F1 to pyruvate, l -lactate, acetate, and propionate, which are detected by the KT2440 receptor, and the medium- and long-chain carboxylic acids hexanoic acid and dodecanoic acid. β-Galactosidase assays of P. putida F1 carrying an mcfP-lacZ transcriptional fusion showed that the mcfP gene is not induced in response to alcohols. Together, our results are consistent with the idea that the carboxylic acids generated from the oxidation of alcohols are the actual attractants sensed by McfP in P. putida F1, rather than the alcohols themselves. IMPORTANCE Alcohols, released as fermentation products and produced as intermediates in the catabolism of many organic compounds, including hydrocarbons and fatty acids, are common components of the microbial food web in soil and sediments. Although they serve as good carbon and energy sources for many soil bacteria, alcohols have primarily been reported to be repellents rather than attractants for motile bacteria. Little is known about how alcohols are sensed by microbes in the environment. We report here that catabolizable n -alcohols with linear chains of up to 12 carbons serve as attractants for the soil bacterium Pseudomonas putida , and rather than being detected directly, alcohols appear to be catabolized to acetate, which is then sensed by a specific cell-surface chemoreceptor protein. 
    more » « less