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Adaptive laboratory evolution of Clostridium autoethanogenum to metabolize CO2 and H2 enhances growth rates in chemostat and unravels proteome and metabolome alterations

ABSTRACT: Gas fermentation of CO₂ and H₂ is an attractive means to sustainably produce fuels and chemicals. Clostridium autoethanogenum is a model organism for industrial CO-to-ethanol and presents an opportunity for CO₂-to-ethanol processes. As we have previously characterized its CO₂/H₂ chemostat growth, here we use adaptive laboratory evolution (ALE) with the aim of improving growth with CO₂/H₂. Seven ALE lineages were generated, all with improved specific growth rates. Developed with 2% CO supplementation of CO₂/H₂, Evolved lineage D has the highest ethanol/acetate of ALE lineages when fermenting CO₂/H₂. Chemostat comparison against the parental strain shows no change in acetate or ethanol production, while Evolved D could achieve a higher maximum dilution rate. Complete multi-omics analyses at steady-state revealed that although Evolved D has widespread proteome changes, intracellular metabolites prevent phenotype shifts. Yet, we observe numerous insights to CO₂/H₂ metabolism via these multi-omics results and link these to mutations, suggesting novel targets for metabolic engineering.

INSTRUMENT(S): Q Exactive HF

ORGANISM(S): Clostridium Autoethanogenum

SUBMITTER: James Heffernan

LAB HEAD: Esteban Marcellin

PROVIDER: PXD048075 | Pride | 2024-05-09

REPOSITORIES: Pride

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Adaptive laboratory evolution of Clostridium autoethanogenum to metabolize CO2 and H2 enhances growth rates in chemostat and unravels proteome and metabolome alterations

Project description:Gas fermentation of CO₂ and H₂ is an attractive means to sustainably produce fuels and chemicals. Clostridium autoethanogenum is a model organism for industrial CO-to-ethanol and presents an opportunity for CO₂-to-ethanol processes. As we have previously characterized its CO₂/H₂ chemostat growth, here we use adaptive laboratory evolution (ALE) with the aim of improving growth with CO₂/H₂. Seven ALE lineages were generated, all with improved specific growth rates. Developed with 2% CO supplementation of CO₂/H₂, Evolved lineage D has the highest ethanol/acetate of ALE lineages when fermenting CO₂/H₂. Chemostat comparison against the parental strain shows no change in acetate or ethanol production, while Evolved D could achieve a higher maximum dilution rate. Complete multi-omics analyses at steady-state revealed that although Evolved D has widespread proteome changes, intracellular metabolites prevent phenotype shifts. Yet, we observe numerous insights to CO₂/H₂ metabolism via these multi-omics results and link these to mutations, suggesting novel targets for metabolic engineering.

2024-05-09 | PXD048143 | Pride

Proteomic profiling of ethanol tolerance in yeast

Project description:For yeast cells, tolerance to high levels of ethanol is vital both in their natural environment and in industrially relevant conditions. We recently genotyped experimentally evolved yeast strains, tolerant to high levels of ethanol. In this study, we elucidate how the genotype and phenotype are related to each other on the molecular level through an integrated analysis of mutation data with protein abundance profiles from six evolved, ethanol-tolerant clones. We constructed a protein network of regulatory proteins, mutated and differentially regulated proteins. This resulted in the identification of a rewired regulatory network after adaptation to high levels of ethanol.

2020-12-03 | PXD006631 | Pride

Proteomic Investigation of Evolved Daptomycin-Tolerant MRSA due to Mutation in YycH

Project description:Several groups have shown that through evolution experiments, tolerance and resistance evolved rapidly under cyclic antibiotic treatment. In other words, intermittent antibiotic exposure performed in a typical adaptive laboratory evolution (ALE) experiments will “train” the bacteria to become tolerant/resistant to the drug. Although ALE has added new knowledge regarding the impact of varying treatment conditions on the evolution of tolerance/resistance, the role of some parameters such as population bottlenecks remains poorly understood. In this study, we employed ALE to investigate the evolution of methicillin-resistant S. aureus under repetitive daptomycin treatment using a modified protocol that incorporated population bottleneck following antibiotic exposure. We observed that although tolerance development is slower under bottlenecking conditions, the populations finally attained tolerance mutation in the yycH gene after twelve cycles of treatment. Extending the evolution experiment and changing the treatment scheme to a fast evolution protocol (treatment during exponential phase without bottlenecking) led to the emergence of daptomycin resistance (mutation in mprF gene). Through proteomics, we uncovered the differential adaptation strategies of these daptomycin tolerant and resistant MRSA strains, and how they respond differently to antibiotics compared to the ancestral wild-type.

2022-07-16 | PXD031078 | Pride

Project description:Pseudomonas putida KT2440 isolates after ALE on ethanol

| PRJNA642834 | ENA

Fuel Ethanol Yeast Strains

Project description:Fuel ethanol is now considered a global energy commodity that is fully competitive with gasoline. We have determined genome copy number differences that are common to five industrially important fuel ethanol yeast strains responsible for the production of billions of gallons of fuel ethanol per year from sugarcane. The fuel strains used were CAT1, BG1, PE2, SA1, and VR1 (note that two independent isolates were analyzed, denoted by "-1" and "-2"). These array-CGH data were compared with array-CGH data from nine other non-fuel industrial yeasts: An ale brewing strain ("Sc-ale"), four wine strains (GSY2A, GSY3A, GSY10A, GSY11B), and 4 bakers' yeast strains (GSY149, GSY150, GSY154, GSY155). Our results reveal significant amplifications of the telomeric SNO and SNZ genes only in the fuel strains, whose protein products are involved in the biosynthesis of vitamins B6 (pyridoxine) and B1 (thiamin). We show that these amplifications allow these yeasts to grow efficiently, especially at high sugar concentrations, regardless of the presence or absence of either of the two vitamins. Our results reveal important genetic adaptations that have been selected for in the industrial environment, which may be required for the efficient fermentation of biomass-derived sugars from other renewable feedstocks. A strain or line experiment design type assays differences between multiple strains, cultivars, serovars, isolates, lines from organisms of a single species. Strain Name: fuel strains used for aCGH Strain_or_line_design

2010-06-25 | E-GEOD-13875 | biostudies-arrayexpress

Escherichia coli strain:BW25113

Project description:Genome sequencing data of evolved strains from ALE experiments

| PRJDB6152 | ENA

Proteome Differences between Daptomycin-Resistant and WT MRSA Biofilms after Elasnin Treatment

Project description:A recent study reported that daptomycin-resistant MRSA (DAPR) strain biofilm is more resistant to daptomycin and vancomycin, as compared to the WT strain biofilm. This pose a great danger since DAPR MRSA is prevalent in clinics and they often form biofilms in medical devices. In this study, we investigate the anti-biofilm activity of elasnin against DAPR MRSA biofilms, and we observed that elasnin is not only effective to eradicate the biofilm of the DAPR strain, but it shows a superior activity compared to the susceptible WT strain. Using proteomics, we compared the proteome profile of the DAPR and the WT strain biofilm cells under elasnin treatment to reveal why elasin is superior against the DAPR strain. Besides, we also employed adaptive laboratory evolution (ALE) experiment by repetitively treating MRSA culture with high dose of elasnin, and generated evolved MRSA strain with increased elasnin tolerance. Using quantitative proteomics, we compared the proteome differences in the elasnin-susceptible WT MRSA and elasnin-tolerant evolved strain.

2021-11-11 | PXD024005 | Pride

MIR sequences recruit zinc finger protein ZNF768 to expressed genes

Project description:Mammalian-wide interspersed repeats (MIRs) are retrotransposed elements of mammalian genomes. Here we report the specific binding of zinc finger protein ZNF768 to the sequence motif GCTGTGTG (N20) CCTCTCTG in the core region of MIRs. ZNF768 binding is preferentially associated with euchromatin and promoter regions of genes. Binding was observed for genes expressed in a cell type-specific manner in human B cell line Raji and osteosarcoma U2OS cells. Mass spectrometric analysis of ZNF768 associated proteins revealed binding of ZNF768 to Elongator subunits 1, 2, and 3. The N-terminus of ZNF768 contains a heptad repeat array structurally related to the C-terminal domain (CTD) of RNA polymerase II. This array evolved in placental animals but not marsupials and monotreme species, displays species-specific length variations, and possibly fulfills CTD related functions in gene regulation. We propose that the evolution of MIRs and ZNF768 has extended the repertoire of gene regulatory mechanisms in mammals and that ZNF768 binding is associated with cell type-specific gene expression.

2018-10-29 | PXD010831 | Pride

Fuel Ethanol Yeast Strains

2009-09-24 | GSE13875 | GEO

Transcriptomic analysis of two ethanol-tolerant Saccharomyces cerevisiae clones obtained by evolutionary engineering

Project description:Evolutionary engineering strategy was used for selection of ethanol-tolerant Saccharomyces cerevisiae clones under gradually increasing ethanol stress levels. Clones B2 and B8 were selected based on their higher ethanol-tolerance and higher ethanol production levels. Whole genome microarray analysis was used for identifying the gene expression levels of these two evolved clones compared to the reference strain.

2016-03-01 | GSE78759 | GEO

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