scholarly journals Clostridium cellulovorans Proteomic Responses to Butanol Stress

2021 ◽  
Vol 12 ◽  
Author(s):  
Paolo Costa ◽  
Giulia Usai ◽  
Angela Re ◽  
Marcello Manfredi ◽  
Giuseppe Mannino ◽  
...  
Keyword(s):  
Protein Quality ◽  
Cell Envelope ◽  
Protein Biosynthesis ◽  
Saturated Fatty Acids ◽  
Atp Synthesis ◽  
Study Data ◽  
Direct Production ◽  
Clostridium Cellulovorans ◽  
Butanol Tolerance ◽  
Microbial Strains

Combination of butanol-hyperproducing and hypertolerant phenotypes is essential for developing microbial strains suitable for industrial production of bio-butanol, one of the most promising liquid biofuels. Clostridium cellulovorans is among the microbial strains with the highest potential for direct production of n-butanol from lignocellulosic wastes, a process that would significantly reduce the cost of bio-butanol. However, butanol exhibits higher toxicity compared to ethanol and C. cellulovorans tolerance to this solvent is low. In the present investigation, comparative gel-free proteomics was used to study the response of C. cellulovorans to butanol challenge and understand the tolerance mechanisms activated in this condition. Sequential Window Acquisition of all Theoretical fragment ion spectra Mass Spectrometry (SWATH-MS) analysis allowed identification and quantification of differentially expressed soluble proteins. The study data are available via ProteomeXchange with the identifier PXD024183. The most important response concerned modulation of protein biosynthesis, folding and degradation. Coherent with previous studies on other bacteria, several heat shock proteins (HSPs), involved in protein quality control, were up-regulated such as the chaperones GroES (Cpn10), Hsp90, and DnaJ. Globally, our data indicate that protein biosynthesis is reduced, likely not to overload HSPs. Several additional metabolic adaptations were triggered by butanol exposure such as the up-regulation of V- and F-type ATPases (involved in ATP synthesis/generation of proton motive force), enzymes involved in amino acid (e.g., arginine, lysine, methionine, and branched chain amino acids) biosynthesis and proteins involved in cell envelope re-arrangement (e.g., the products of Clocel_4136, Clocel_4137, Clocel_4144, Clocel_4162 and Clocel_4352, involved in the biosynthesis of saturated fatty acids) and a redistribution of carbon flux through fermentative pathways (acetate and formate yields were increased and decreased, respectively). Based on these experimental findings, several potential gene targets for metabolic engineering strategies aimed at improving butanol tolerance in C. cellulovorans are suggested. This includes overexpression of HSPs (e.g., GroES, Hsp90, DnaJ, ClpC), RNA chaperone Hfq, V- and F-type ATPases and a number of genes whose function in C. cellulovorans is currently unknown.

scholarly journals Proteomic Signatures of Clostridium difficile Stressed with Metronidazole, Vancomycin, or Fidaxomicin

Cells ◽  
2018 ◽  
Vol 7 (11) ◽  
pp. 213 ◽  
Author(s):  
Sandra Maaß ◽  
Andreas Otto ◽  
Dirk Albrecht ◽  
Katharina Riedel ◽  
Anke Trautwein-Schult ◽  
...  
Keyword(s):  
Clostridium Difficile ◽  
Antibiotic Treatment ◽  
Antimicrobial Agents ◽  
Cell Envelope ◽  
Protein Biosynthesis ◽  
Cellular Functions ◽  
Antibiotic Resistant ◽  
Incidence And Mortality ◽  
A Cell ◽  
Proteomic Response

The anaerobic pathogen Clostridium difficile is of growing significance for the health care system due to its increasing incidence and mortality. As C. difficile infection is both supported and treated by antibiotics, a deeper knowledge on how antimicrobial agents affect the physiology of this important pathogen may help to understand and prevent the development and spreading of antibiotic resistant strains. As the proteomic response of a cell to stress aims at counteracting the harmful effects of this stress, it can be expected that the pattern of a pathogen’s responses to antibiotic treatment will be dependent on the antibiotic mechanism of action. Hence, every antibiotic treatment is expected to result in a specific proteomic signature characterizing its mode of action. In the study presented here, the proteomic response of C. difficile 630∆erm to vancomycin, metronidazole, and fidaxomicin stress was investigated on the level of protein abundance and protein synthesis based on 2D PAGE. The quantification of 425 proteins of C. difficile allowed the deduction of proteomic signatures specific for each drug treatment. Indeed, these proteomic signatures indicate very specific cellular responses to each antibiotic with only little overlap of the responses. Whereas signature proteins for vancomycin stress fulfil various cellular functions, the proteomic signature of metronidazole stress is characterized by alterations of proteins involved in protein biosynthesis and protein degradation as well as in DNA replication, recombination, and repair. In contrast, proteins differentially expressed after fidaxomicin treatment can be assigned to amino acid biosynthesis, transcription, cell motility, and the cell envelope functions. Notably, the data provided by this study hint also at so far unknown antibiotic detoxification mechanisms.

scholarly journals Multiple pathways of toxicity induced by C9orf72 dipeptide repeat aggregates and G4C2 RNA in a cellular model

2020 ◽  
Author(s):  
Frédéric Frottin ◽  
Manuela Pérez-Berlanga ◽  
F. Ulrich Hartl ◽  
Mark S. Hipp
Keyword(s):  
Protein Transport ◽  
Protein Quality ◽  
Protein Biosynthesis ◽  
Abundant Species ◽  
Cellular Model ◽  
Protein Biogenesis ◽  
Pathways Of Toxicity ◽  
Lateral Sclerosis ◽  
Dipeptide Repeat ◽  
Pronounced Inhibition

AbstractThe most frequent genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia is a G4C2 repeat expansion in the C9orf72 gene. This expansion gives rise to translation of aggregating dipeptide repeat (DPR) proteins, including poly-GA as the most abundant species. However, gain of toxic function effects have been attributed to either the DPRs or the pathological G4C2 RNA. Here we analyzed in a cellular model the relative toxicity of DPRs and RNA. Cytoplasmic poly-GA aggregates, generated in the absence of G4C2 RNA, interfered with nucleocytoplasmic protein transport, but had little effect on cell viability. In contrast, nuclear poly-GA was more toxic, impairing nucleolar protein quality control and protein biosynthesis. Production of the G4C2 RNA strongly reduced viability independent of DPR translation and caused pronounced inhibition of nuclear mRNA export and protein biogenesis. Thus, while the toxic effects of G4C2 RNA predominate, DPRs exert additive effects that may contribute to pathology.

scholarly journals Functional compartmentalization and metabolic separation in a prokaryotic cell

2021 ◽  
Vol 118 (25) ◽  
pp. e2022114118
Author(s):  
Jennifer Flechsler ◽  
Thomas Heimerl ◽  
Harald Huber ◽  
Reinhard Rachel ◽  
Ivan A. Berg
Keyword(s):  
Outer Membrane ◽  
Protein Biosynthesis ◽  
Atp Synthesis ◽  
Side Reactions ◽  
Strictly Anaerobic ◽  
Prokaryotic Cell ◽  
Dna And Rna ◽  
Information Processes ◽  
Cytoplasmic Compartment ◽  
Gradient Generation

The prokaryotic cell is traditionally seen as a “bag of enzymes,” yet its organization is much more complex than in this simplified view. By now, various microcompartments encapsulating metabolic enzymes or pathways are known for Bacteria. These microcompartments are usually small, encapsulating and concentrating only a few enzymes, thus protecting the cell from toxic intermediates or preventing unwanted side reactions. The hyperthermophilic, strictly anaerobic Crenarchaeon Ignicoccus hospitalis is an extraordinary organism possessing two membranes, an inner and an energized outer membrane. The outer membrane (termed here outer cytoplasmic membrane) harbors enzymes involved in proton gradient generation and ATP synthesis. These two membranes are separated by an intermembrane compartment, whose function is unknown. Major information processes like DNA replication, RNA synthesis, and protein biosynthesis are located inside the “cytoplasm” or central cytoplasmic compartment. Here, we show by immunogold labeling of ultrathin sections that enzymes involved in autotrophic CO2 assimilation are located in the intermembrane compartment that we name (now) a peripheric cytoplasmic compartment. This separation may protect DNA and RNA from reactive aldehydes arising in the I. hospitalis carbon metabolism. This compartmentalization of metabolic pathways and information processes is unprecedented in the prokaryotic world, representing a unique example of spatiofunctional compartmentalization in the second domain of life.

Cell envelope proteases and peptidases of Pseudomonas aeruginosa: multiple roles, multiple mechanisms

2020 ◽  
Vol 44 (6) ◽  
pp. 857-873
Author(s):  
Astra Heywood ◽  
Iain L Lamont
Keyword(s):  
Pseudomonas Aeruginosa ◽  
Outer Membrane ◽  
Protein Quality ◽  
Environmental Changes ◽  
Cell Envelope ◽  
Opportunistic Pathogen ◽  
Multiple Roles ◽  
Current State ◽  
Wide Range ◽  
Membrane Cell

ABSTRACT Pseudomonas aeruginosa is a Gram-negative bacterium that is commonly isolated from damp environments. It is also a major opportunistic pathogen, causing a wide range of problematic infections. The cell envelope of P. aeruginosa, comprising the cytoplasmic membrane, periplasmic space, peptidoglycan layer and outer membrane, is critical to the bacteria's ability to adapt and thrive in a wide range of environments. Over 40 proteases and peptidases are located in the P. aeruginosa cell envelope. These enzymes play many crucial roles. They are required for protein secretion out of the cytoplasm to the periplasm, outer membrane, cell surface or the environment; for protein quality control and removal of misfolded proteins; for controlling gene expression, allowing adaptation to environmental changes; for modification and remodelling of peptidoglycan; and for metabolism of small molecules. The key roles of cell envelope proteases in ensuring normal cell functioning have prompted the development of inhibitors targeting some of these enzymes as potential new anti-Pseudomonas therapies. In this review, we summarise the current state of knowledge across the breadth of P. aeruginosa cell envelope proteases and peptidases, with an emphasis on recent findings, and highlight likely future directions in their study.

254 MOLECULAR AND SUBCELLULAR CHARACTERIZATION OF OOCYTES SCREENED FOR THEIR DEVELOPMENTAL COMPETENCE BASED ON G6PDH ACTIVITY

2008 ◽  
Vol 20 (1) ◽  
pp. 207
Author(s):  
H. Torner ◽  
N. Ghanem ◽  
C. Ambros ◽  
M. Hoelker ◽  
W. Tomek ◽  
...  
Keyword(s):  
Map Kinase ◽  
Protein Biosynthesis ◽  
Expression Profiles ◽  
Gene Expression Profiles ◽  
Atp Synthesis ◽  
Developmental Competence ◽  
Mitochondrial Activity ◽  
G6pdh Activity ◽  
Cresyl Blue

Oocyte selection based on glucose-6-phosphate dehydrogenase (G6PDH) activity has been successfully used to differentiate between competent and incompetent bovine oocytes (Alm 2005 Theriogenology 63, 2194–2205). However, the intrinsic molecular and subcellular characteristics of these oocytes have not yet been investigated. Here we aim to compare the developmental, molecular, and subcellular characteristics of oocytes selected using brilliant cresyl blue (BCB) staining based on G6PDH activity. Immature compact cumulus–oocyte complexes (COCs) were stained with 26 µm BCB (B-5388, Sigma-Alderich, Taufenkirchen, Germany) for 90 min. Based on their coloration, oocytes were divided into BCB– (colorless cytoplasm, high G6PDH activity) and BCB+ (colored cytoplasm, low G6PDH activity). The chromatin configuration and the mitochondrial activity of oocytes were determined by fluorescence labelling and photometric measurement (n = 337). The abundance and phosphorylation pattern of protein kinases Akt and MAP kinase were estimated by western blot analysis (n = 500). A bovine cDNA microarray with 2000 clones was used to analyze the gene expression profiles of BCB+ and BCB– oocytes (n = 580). BCB+ oocytes were found to result in a higher blastocyst rate (33.1 � 3.1%) until Day 8 of in vitro culture compared to BCB– ones (12.1 � 1.5%). Moreover, BCB+ oocytes showed higher phosphorylation levels of Akt and MAP kinase compared to the BCB– oocytes. After array data analysis, BCB+ oocytes were found to be enriched with genes regulating transcription (SMARCA5), cell cycle (NASP), and protein biosynthesis (RPS274A and EEF1A1), while the BCB– oocytes had a higher level of genes involved in ATP synthesis (ATP5A1), mitochondrial electron transport (FL405), calcium ion binding (S100A10), and growth factor activity (BMP-15). Independent real-time quantitative PCR validated 90% (9/10) of the genes investigated to be in agreement with the array expression profile. The study has shown evidence of differences in molecular and subcellular organization of oocytes with different G6PDH activity.

scholarly journals Epigenetic Regulation of Verticillium dahliae Virulence: Does DNA Methylation Level Play A Role?

2020 ◽  
Vol 21 (15) ◽  
pp. 5197
Author(s):  
Jorge A. Ramírez-Tejero ◽  
Carmen Gómez-Lama Cabanás ◽  
Antonio Valverde-Corredor ◽  
Jesús Mercado-Blanco ◽  
Francisco Luque
Keyword(s):  
Dna Methylation ◽  
Verticillium Dahliae ◽  
Protein Biosynthesis ◽  
Chromatin Condensation ◽  
Inverse Correlation ◽  
Atp Synthesis ◽  
Infection Process ◽  
Epigenetic Mechanism ◽  
High Plasticity ◽  
Protein Coding

Verticillium dahliae is the etiological agent of Verticillium wilt of olive. The virulence of Defoliating V. dahliae isolates usually displays differences and high plasticity. This work studied whether an epigenetic mechanism was involved in this plasticity. An inverse correlation between virulence and DNA methylation of protein-coding genes was found. A set of 831 genes was selected for their highly consistent inverse methylation profile and virulence in the five studied isolates. Of these genes, ATP-synthesis was highly represented, which indicates that the more virulent D isolates are, the more energy requirements they may have. Furthermore, there were numerous genes in the protein biosynthesis process: genes coding for the chromatin structure, which suggests that epigenetic changes may also affect chromatin condensation; many transmembrane transporter genes, which is consistent with denser compounds, traffic through membranes in more virulent isolates; a fucose-specific lectin that may play a role in the attachment to plant cell walls during the host infection process; and pathogenic cutinases that facilitate plant invasion and sporulation genes for rapid spreading alongside plants. Our findings support the notion that differences in the virulence of the Defoliating V. dahliae isolates may be controlled, at least to some extent, by an epigenetic mechanism.

scholarly journals The transcriptional regulator of the chaperone response HSF1 controls hepatic bioenergetics and protein homeostasis

2017 ◽  
Vol 216 (3) ◽  
pp. 723-741 ◽  
Author(s):  
Aijun Qiao ◽  
Xiongjie Jin ◽  
Junfeng Pang ◽  
Demetrius Moskophidis ◽  
Nahid F. Mivechi
Keyword(s):  
Transcriptional Repression ◽  
Protein Quality ◽  
Metabolic Diseases ◽  
Atp Synthesis ◽  
Transcriptional Regulator ◽  
Metabolic Energy ◽  
Metabolic Adaptation ◽  
Protein Homeostasis ◽  
Heat Shock Factor 1 ◽  
Nicotinamide Phosphoribosyltransferase

Metabolic energy reprogramming facilitates adaptations to a variety of stress conditions and cellular dysfunction, but how the energetic demands are monitored and met in response to physiological stimuli remains elusive. Our data support a model demonstrating that heat shock factor 1 (HSF1), a master transcriptional regulator of the chaperone response, has been coopted from its role as a critical protein quality-control regulator to having a central role in systemic energy sensing and for metabolic adaptation to nutrient availability. We found that in the absence of HSF1, levels of NAD+ and ATP are not efficiently sustained in hepatic cells, largely because of transcriptional repression of nicotinamide phosphoribosyltransferase in the NAD+ salvage pathway. Mechanistically, the defect in NAD+ and ATP synthesis linked to a loss of NAD+-dependent deacetylase activity, increased protein acetylation, and impaired mitochondrial integrity. Remarkably, the drop in ATP level caused by HSF1 loss invoked an adaptive response featuring the inhibition of energetically demanding processes, including gluconeogenesis, translation, and lipid synthesis. Our work identifies HSF1 as a central regulator of cellular bioenergetics and protein homeostasis that benefits malignant cell progression and exacerbates development of metabolic diseases.

scholarly journals Localized Proteotoxic Stress in Mitochondrial Intermembrane Space and Matrix Elicits Sub-compartment Specific Response Pathways Governed by Unique Modulators

2020 ◽  
Author(s):  
Kannan Boosi Narayana Rao ◽  
Pratima Pandey ◽  
Rajasri Sarkar ◽  
Asmita Ghosh ◽  
Shemin Mansuri ◽  
...  
Keyword(s):  
Stress Response ◽  
Cellular Response ◽  
Protein Quality ◽  
Atp Synthesis ◽  
Specific Response ◽  
Intermembrane Space ◽  
Matrix Stress ◽  
Proteotoxic Stress ◽  
Tom Complex ◽  
The Matrix

AbstractThe complex double-membrane architecture of mitochondria is essential for its ATP synthesis function and divides the organelle into two sub-mitochondrial compartments, inter-membrane space (IMS) and matrix. The folding environments of IMS and matrix are significantly different owing to its dissimilar oxido-reductive environments and distinctly divergent protein quality control (PQC) machineries. Here, by inducing proteotoxic stress restricted to IMS or matrix by targeting three different stressor proteins, we show that the cellular response to IMS or matrix-localized misfolding stress is distinct and unique. IMS and matrix stress response pathways are quite effective in combatting stress despite significant stress-induced alteration in mitochondrial phenotypes. IMS misfolding stress leads to specific upregulation of IMS chaperones and components of TOM complex while matrix chaperones and cytosolic PQC components are upregulated during matrix stress. Notably, the amplitude of upregulation of mitochondrial chaperones is not overwhelming. We report that cells respond to mitochondrial stress through an adaptive mechanism by adjourning mitochondrial respiration while upregulating glycolysis as a compensatory pathway. We show that subunits of TOM complex act as specific modulators of IMS-stress response while Vms1 precisely modulates the matrix stress response.

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