scholarly journals The TOR kinase pathway is relevant for nitrogen signaling and antagonism of the mycoparasite Trichoderma atroviride

PLoS ONE ◽  
2021 ◽  
Vol 16 (12) ◽  
pp. e0262180
Author(s):  
Rossana Segreto ◽  
Hoda Bazafkan ◽  
Julia Millinger ◽  
Martina Schenk ◽  
Lea Atanasova ◽  
...  
Keyword(s):  
Secondary Metabolites ◽  
Mammalian Cells ◽  
Ribosome Biogenesis ◽  
Genetic Manipulation ◽  
Cellular Growth ◽  
Glycoside Hydrolases ◽  
Yeast Genome ◽  
Minor Effect ◽  
Trichoderma Atroviride ◽  
Tor Kinase

Trichoderma atroviride (Ascomycota, Sordariomycetes) is a well-known mycoparasite applied for protecting plants against fungal pathogens. Its mycoparasitic activity involves processes shared with plant and human pathogenic fungi such as the production of cell wall degrading enzymes and secondary metabolites and is tightly regulated by environmental cues. In eukaryotes, the conserved Target of Rapamycin (TOR) kinase serves as a central regulator of cellular growth in response to nutrient availability. Here we describe how alteration of the activity of TOR1, the single and essential TOR kinase of T. atroviride, by treatment with chemical TOR inhibitors or by genetic manipulation of selected TOR pathway components affected various cellular functions. Loss of TSC1 and TSC2, that are negative regulators of TOR complex 1 (TORC1) in mammalian cells, resulted in altered nitrogen source-dependent growth of T. atroviride, reduced mycoparasitic overgrowth and, in the case of Δtsc1, a diminished production of numerous secondary metabolites. Deletion of the gene encoding the GTPase RHE2, whose mammalian orthologue activates mTORC1, led to rapamycin hypersensitivity and altered secondary metabolism, but had an only minor effect on vegetative growth and mycoparasitic overgrowth. The latter also applied to mutants missing the npr1-1 gene that encodes a fungus-specific kinase known as TOR target in yeast. Genome-wide transcriptome analysis confirmed TOR1 as a regulatory hub that governs T. atroviride metabolism and processes associated to ribosome biogenesis, gene expression and translation. In addition, mycoparasitism-relevant genes encoding terpenoid and polyketide synthases, peptidases, glycoside hydrolases, small secreted cysteine-rich proteins, and G protein coupled receptors emerged as TOR1 targets. Our results provide the first in-depth insights into TOR signaling in a fungal mycoparasite and emphasize its importance in the regulation of processes that critically contribute to the antagonistic activity of T. atroviride.

scholarly journals Commuting to Work: Nucleolar Long Non-Coding RNA Control Ribosome Biogenesis from Near and Far

2021 ◽  
Vol 7 (3) ◽  
pp. 42
Author(s):  
Victoria Mamontova ◽  
Barbara Trifault ◽  
Lea Boten ◽  
Kaspar Burger
Keyword(s):  
Gene Expression ◽  
Rna Polymerase Ii ◽  
Ribosome Biogenesis ◽  
Intergenic Spacer ◽  
Cellular Growth ◽  
Rrna Synthesis ◽  
Protein Coding ◽  
Non Coding Rna ◽  
And Function ◽  
In Cis

Gene expression is an essential process for cellular growth, proliferation, and differentiation. The transcription of protein-coding genes and non-coding loci depends on RNA polymerases. Interestingly, numerous loci encode long non-coding (lnc)RNA transcripts that are transcribed by RNA polymerase II (RNAPII) and fine-tune the RNA metabolism. The nucleolus is a prime example of how different lncRNA species concomitantly regulate gene expression by facilitating the production and processing of ribosomal (r)RNA for ribosome biogenesis. Here, we summarise the current findings on how RNAPII influences nucleolar structure and function. We describe how RNAPII-dependent lncRNA can both promote nucleolar integrity and inhibit ribosomal (r)RNA synthesis by modulating the availability of rRNA synthesis factors in trans. Surprisingly, some lncRNA transcripts can directly originate from nucleolar loci and function in cis. The nucleolar intergenic spacer (IGS), for example, encodes nucleolar transcripts that counteract spurious rRNA synthesis in unperturbed cells. In response to DNA damage, RNAPII-dependent lncRNA originates directly at broken ribosomal (r)DNA loci and is processed into small ncRNA, possibly to modulate DNA repair. Thus, lncRNA-mediated regulation of nucleolar biology occurs by several modes of action and is more direct than anticipated, pointing to an intimate crosstalk of RNA metabolic events.

scholarly journals The Small Subunit Processome Is Required for Cell Cycle Progression at G1

2004 ◽  
Vol 15 (11) ◽  
pp. 5038-5046 ◽  
Author(s):  
Kara A. Bernstein ◽  
Susan J. Baserga
Keyword(s):  
Cell Cycle ◽  
Ribosome Biogenesis ◽  
Cell Cycle Progression ◽  
Small Subunit ◽  
Cellular Growth ◽  
Yeast Cells ◽  
G1 Phase ◽  
Relay Cell ◽  
Ssu Processome ◽  
Nucleolar Rna

Without ribosome biogenesis, translation of mRNA into protein ceases and cellular growth stops. We asked whether ribosome biogenesis is cell cycle regulated in the yeasts Saccharomyces cerevisiae and Schizosaccharomyces pombe, and we determined that it is not regulated in the same manner as in metazoan cells. We therefore turned our attention to cellular sensors that relay cell size information via ribosome biogenesis. Our results indicate that the small subunit (SSU) processome, a complex consisting of 40 proteins and the U3 small nucleolar RNA necessary for ribosome biogenesis, is not mitotically regulated. Furthermore, Nan1/Utp17, an SSU processome protein, does not provide a link between ribosome biogenesis and cell growth. However, when individual SSU processome proteins are depleted, cells arrest in the G1 phase of the cell cycle. This arrest was further supported by the lack of staining for proteins expressed in post-G1. Similarly, synchronized cells depleted of SSU processome proteins did not enter G2. This suggests that when ribosomes are no longer made, the cells stall in the G1. Therefore, yeast cells must grow to a critical size, which is dependent upon having a sufficient number of ribosomes during the G1 phase of the cell cycle, before cell division can occur.

scholarly journals Changes in Membrane Fluid State and Heat Shock Response Cause Attenuation of Virulence

2010 ◽  
Vol 192 (7) ◽  
pp. 1999-2005 ◽  
Author(s):  
Amalia Porta ◽  
Annamaria Eletto ◽  
Zsolt Török ◽  
Silvia Franceschelli ◽  
Attila Glatz ◽  
...  
Keyword(s):  
Heat Shock ◽  
Genetic Modification ◽  
Heat Shock Response ◽  
Mammalian Cells ◽  
Histoplasma Capsulatum ◽  
Genetic Manipulation ◽  
Microbial Pathogens ◽  
Fluid State ◽  
Shock Response ◽  
Serovar Typhimurium

ABSTRACT So far attenuation of pathogens has been mainly obtained by chemical or heat treatment of microbial pathogens. Recently, live attenuated strains have been produced by genetic modification. We have previously demonstrated that in several prokaryotes as well as in yeasts and mammalian cells the heat shock response is controlled by the membrane physical state (MPS). We have also shown that in Salmonella enterica serovar Typhimurium LT2 (Salmonella Typhimurium) overexpression of a Δ12-desaturase gene alters the MPS, inducing a sharp impairment of transcription of major heat shock genes and failure of the pathogen to grow inside macrophage (MΦ) (A. Porta et al., J. Bacteriol. 192:1988-1998, 2010). Here, we show that overexpression of a homologous Δ9-desaturase sequence in the highly virulent G217B strain of the human fungal pathogen Histoplasma capsulatum causes loss of its ability to survive and persist within murine MΦ along with the impairment of the heat shock response. When the attenuated strain of H. capsulatum was injected in a mouse model of infection, it did not cause disease. Further, treated mice were protected when challenged with the virulent fungal parental strain. Attenuation of virulence in MΦ of two evolutionarily distant pathogens was obtained by genetic modification of the MPS, suggesting that this is a new method that may be used to produce attenuation or loss of virulence in both other intracellular prokaryotic and eukaryotic pathogens. This new procedure to generate attenuated forms of pathogens may be used eventually to produce a novel class of vaccines based on the genetic manipulation of a pathogen's membrane fluid state and stress response.

scholarly journals Mushroom Biomass: Some Clinical Implications of β-Glucans and Enzymes

2016 ◽  
Vol 4 (Special-Issue-October) ◽  
pp. 37-47
Author(s):  
Ana Barros ◽  
Vitoria Bell ◽  
Jorge Ferrão ◽  
Vittorio Calabrese ◽  
Tito Fernandes
Keyword(s):  
Secondary Metabolites ◽  
Dietary Supplement ◽  
Biological Response ◽  
Cellular Growth ◽  
Health Promoting ◽  
The Rich ◽  
The Difference ◽  
Cytochrome P 450

Mushrooms have attracted market attention because they are a potential source of bioactive compounds able to perform several functions in organisms with benefits for the health of the consumer. Cultivation processes vary according a) industrial fermentation - in large vats to produce extracted form of mushrooms or b) closed cultivation system - individually grown in jars on an aseptic “substrate” with controlled lighting and irrigation to produce a biomass form of mushrooms. Biomass is the mycelium with primordia (young fruiting body - before the mushroom blooms) containing all the nutrients and active compounds, including β-glucans, enzymes and secondary metabolites. The classification of mushroom biomass varies according to the presentation; the biomass can be classified as a “food” if in powder form or, classified as a “dietary supplement” in tablet form. While tablet mushroom biomass is considered a dietary supplement, mushroom extracts are designated pharmaceutical compounds, pharmanutrients or nutraceuticals. Here we illustrate the difference between mushrooms in the biomass and extract forms, the similarities and differences on its content on enzymes, secondary metabolites and on β-glucans, as a soluble and fermentable fibre. Of particular note is the rich enzyme activity in the biomass form of mushrooms. Such activity includes enzymes that prevent oxidative stress (superoxide dismutase), enzymes that prevent cellular growth (protease, glucoamylase) and enzymes that promote detoxification (cytochrome P-450, peroxidase, glucose-2-oxidase). β-glucans have been proposed to act as “biological response modifiers” based on their effects on the immune system, and its role in the prevention and treatment of various metabolic syndrome-linked diseases. This review focuses also on some described health-promoting potential of mushroom biomass, all through immunomodulation. The role of intestinal microbiota is enhanced.

scholarly journals Control of ribosomal protein synthesis by Microprocessor complex

2020 ◽  
Author(s):  
Xuan Jiang ◽  
Amit Prabhakar ◽  
Stephanie M. Van der Voorn ◽  
Prajakta Ghatpande ◽  
Barbara Celona ◽  
...  
Keyword(s):  
Cell Growth ◽  
Nuclear Export ◽  
Ribosomal Proteins ◽  
Ribosome Biogenesis ◽  
Cellular Growth ◽  
New Paradigm ◽  
Erythroid Progenitors ◽  
Conditional Deletion ◽  
Microprocessor Complex ◽  
Ribosomal Protein Synthesis

AbstractRibosome biogenesis in eukaryotes requires stoichiometric production and assembly of 80 ribosomal proteins (RPs) and 4 ribosomal RNAs, and its rate must be coordinated with cellular growth. The indispensable regulator of RP biosynthesis is the 5’-terminal oligopyrimidine (TOP) motif, spanning the transcription start site of all RP genes. Here we show that the Microprocessor complex, previously linked to the first step of processing microRNAs (miRNAs), coregulates RP expression by binding the TOP motif of nascent RP mRNAs and stimulating transcription elongation via resolution of DNA/RNA hybrids. Cell growth arrest triggers nuclear export and degradation of the Microprocessor protein Drosha by the E3 ubiquitin ligase Nedd4, accumulation of DNA/RNA hybrids at RP gene loci, decreased RP synthesis, and ribosome deficiency, hence synchronizing ribosome production with cell growth. Conditional deletion of Drosha in erythroid progenitors phenocopies human ribosomopathies, in which ribosomal insufficiency leads to anemia. Outlining a miRNA-independent role of the Microprocessor complex at the interphase between cell growth and ribosome biogenesis offers a new paradigm by which cells alter their protein biosynthetic capacity and cellular metabolism.

scholarly journals Homeostasis of branched-chain amino acids is critical for the activity of TOR signaling in Arabidopsis

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Pengfei Cao ◽  
Sang-Jin Kim ◽  
Anqi Xing ◽  
Craig A Schenck ◽  
Lu Liu ◽  
...  
Keyword(s):  
Amino Acids ◽  
Mammalian Cells ◽  
Branched Chain Amino Acids ◽  
Nutrient Sensing ◽  
Tor Signaling ◽  
Functional Connection ◽  
Primary Input ◽  
Tor Kinase ◽  
Arabidopsis Mutant ◽  
Branched Chain

The target of rapamycin (TOR) kinase is an evolutionarily conserved hub of nutrient sensing and metabolic signaling. In plants, a functional connection of TOR activation with glucose availability was demonstrated, while it is yet unclear whether branched-chain amino acids (BCAAs) are a primary input of TOR signaling as they are in yeast and mammalian cells. Here, we report on the characterization of an Arabidopsis mutant over-accumulating BCAAs. Through chemical interventions targeting TOR and by examining mutants of BCAA biosynthesis and TOR signaling, we found that BCAA over-accumulation leads to up-regulation of TOR activity, which causes reorganization of the actin cytoskeleton and actin-associated endomembranes. Finally, we show that activation of TOR is concomitant with alteration of cell expansion, proliferation and specialized metabolism, leading to pleiotropic effects on plant growth and development. These results demonstrate that BCAAs contribute to plant TOR activation and reveal previously uncharted downstream subcellular processes of TOR signaling.

Co-culture of Trichoderma atroviride SG3403 and Bacillus subtilis 22 improves the production of antifungal secondary metabolites

2020 ◽  
Vol 140 ◽  
pp. 104122 ◽  
Author(s):  
Tingting Li ◽  
Jiaquan Tang ◽  
Valliappan Karuppiah ◽  
Yaqian Li ◽  
Nan Xu ◽  
...  
Keyword(s):  
Bacillus Subtilis ◽  
Secondary Metabolites ◽  
Trichoderma Atroviride

New insights of CRISPR technology in human pathogenic fungi

2019 ◽  
Vol 14 (14) ◽  
pp. 1243-1255 ◽  
Author(s):  
Elvira Román ◽  
Daniel Prieto ◽  
Rebeca Alonso-Monge ◽  
Jesús Pla
Keyword(s):  
Mammalian Cells ◽  
Genetic Manipulation ◽  
Pathogenic Fungi ◽  
Biological Research ◽  
Therapeutic Approaches ◽  
New Therapeutic Approaches ◽  
Short Palindromic Repeat ◽  
Future Outcomes ◽  
Near Future ◽  
Genome Manipulation

Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)-Cas systems have emerged as a powerful tool for genome manipulation. Class 2 type II CRISPR/ CAS9 is so far the most studied system and has been implemented in many biological systems such as mammalian cells, plants, fungi and bacteria. Fungi are important causes of human diseases worldwide. Genetic manipulation of pathogenic fungi is critical to develop new therapeutic approaches and novel antifungals. We will review here the progress done with CRISPR/ CAS9 systems in human pathogenic fungi, with emphasis in Candida albicans and the main modifications that have improved their usefulness in biological research. We finally discuss possible future outcomes and applications to the developed in a near future.

scholarly journals The Health Promoting Bioactivities of Lactuca sativa can be Enhanced by Genetic Modulation of Plant Secondary Metabolites

Metabolites ◽  
2019 ◽  
Vol 9 (5) ◽  
pp. 97 ◽  
Author(s):  
Hammad Ismail ◽  
Anna L. Gillespie ◽  
Danielle Calderwood ◽  
Haroon Iqbal ◽  
Colene Gallagher ◽  
...  
Keyword(s):  
Secondary Metabolites ◽  
Lactuca Sativa ◽  
Genetic Manipulation ◽  
Sesquiterpene Lactones ◽  
Plant Secondary Metabolites ◽  
Rol Genes ◽  
In Vitro Bioactivity ◽  
New Paradigm ◽  
Plant Metabolites

Plant secondary metabolites are protective dietary constituents and rol genes evidently increase the synthesis of these versatile phytochemicals. This study subjected a globally important vegetable, lettuce (Lactuca sativa) to a combination of untargeted metabolomics (LC-QTof-MS) and in vitro bioactivity assays. Specifically, we examined the differences between untransformed cultured lettuce (UnT), lettuce transformed with either rolABC (RA) or rolC (RC) and commercially grown (COM) lettuce. Of the 5333 metabolite features aligned, deconvoluted and quantified 3637, 1792 and 3737 significantly differed in RA, RC and COM, respectively, compared with UnT. In all cases the number of downregulated metabolites exceeded the number increased. In vitro bioactivity assays showed that RA and RC (but not COM) significantly improved the ability of L. sativa to inhibit α-glucosidase, inhibit dipeptidyl peptidase-4 (DPP-4) and stimulate GLP-1 secretion. We putatively identified 76 lettuce metabolites (sesquiterpene lactones, non-phenolic and phenolic compounds) some of which were altered by several thousand percent in RA and RC. Ferulic acid levels increased 3033–9777%, aminooxononanoic acid increased 1141–1803% and 2,3,5,4′tetrahydroxystilbene-2-O-β-d-glucoside increased 40,272–48,008%. Compound activities were confirmed using commercially obtained standards. In conclusion, rol gene transformation significantly alters the metabolome of L.sativa and enhances its antidiabetic properties. There is considerable potential to exploit rol genes to modulate secondary metabolite production for the development of novel functional foods. This investigation serves as a new paradigm whereby genetic manipulation, metabolomic analysis and bioactivity techniques can be combined to enable the discovery of novel natural bioactives and determine the functional significance of plant metabolites.

scholarly journals PCB11 Metabolite, 3,3’-Dichlorobiphenyl-4-ol, Exposure Alters the Expression of Genes Governing Fatty Acid Metabolism in the Absence of Functional Sirtuin 3: Examining the Contribution of MnSOD

Antioxidants ◽  
2018 ◽  
Vol 7 (9) ◽  
pp. 121 ◽  
Author(s):  
Sinthia Alam ◽  
Gwendolyn Carter ◽  
Kimberly Krager ◽  
Xueshu Li ◽  
Hans-Joachim Lehmler ◽  
...  
Keyword(s):  
Fatty Acid ◽  
Fatty Acid Metabolism ◽  
Mitochondrial Respiration ◽  
Mammalian Cells ◽  
Acid Metabolism ◽  
Cellular Growth ◽  
Acid Oxidation ◽  
Sirtuin 3 ◽  
Endogenous Fatty Acid ◽  
Mnsod Activity

Although the production of polychlorinated biphenyls (PCBs) is prohibited, the inadvertent production of certain lower-chlorinated PCB congeners still threatens human health. We and others have identified 3,3’-dichlorobiphenyl (PCB11) and its metabolite, 3,3’-dichlorobiphenyl-4-ol (4OH-PCB11), in human blood, and there is a correlation between exposure to this metabolite and mitochondrial oxidative stress in mammalian cells. Here, we evaluated the downstream effects of 4OH-PCB11 on mitochondrial metabolism and function in the presence and absence of functional Sirtuin 3 (SIRT3), a mitochondrial fidelity protein that protects redox homeostasis. A 24 h exposure to 3 μM 4OH-PCB11 significantly decreased the cellular growth and mitochondrial membrane potential of SIRT3-knockout mouse embryonic fibroblasts (MEFs). Only wild-type cells demonstrated an increase in Manganese superoxide dismutase (MnSOD) activity in response to 4OH-PCB11–induced oxidative injury. This suggests the presence of a SIRT3-mediated post-translational modification to MnSOD, which was impaired in SIRT3-knockout MEFs, which counters the PCB insult. We found that 4OH-PCB11 increased mitochondrial respiration and endogenous fatty-acid oxidation-associated oxygen consumption in SIRT3-knockout MEFs; this appeared to occur because the cells exhausted their reserve respiratory capacity. To determine whether these changes in mitochondrial respiration were accompanied by similar changes in the regulation of fatty acid metabolism, we performed quantitative real-time polymerase chain reaction (qRT-PCR) after a 24 h treatment with 4OH-PCB11. In SIRT3-knockout MEFs, 4OH-PCB11 significantly increased the expression of ten genes controlling fatty acid biosynthesis, metabolism, and transport. When we overexpressed MnSOD in these cells, the expression of six of these genes returned to the baseline level, suggesting that the protective role of SIRT3 against 4OH-PCB11 is partially governed by MnSOD activity.

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