scholarly journals Isolation and Characterization of a Symbiosis-Regulated ras from the Ectomycorrhizal Fungus Laccaria bicolor

2001 ◽  
Vol 14 (5) ◽  
pp. 618-628 ◽  
Author(s):  
S. Sundaram ◽  
S. J. Kim ◽  
H. Suzuki ◽  
C. J. Mcquattie ◽  
S. T. Hiremath ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Morphological Changes ◽  
Single Copy ◽  
Ectomycorrhizal Fungus ◽  
Northern Analysis ◽  
Laccaria Bicolor ◽  
Symbiotic Interaction ◽  
Isolation And Characterization ◽  
Wide Range ◽  
Ras Family

Ectomycorrhizae formed by the symbiotic interaction between ectomycorrhizal fungi and plant roots play a key role in maintaining and improving the health of a wide range of plants. Mycorrhizal initiation, development, and functional maintenance involve morphological changes that are mediated by activation and suppression of several fungal and plant genes. We identified a gene, Lbras, in the ectomycorrhizal fungus Laccaria bicolor that belongs to the ras family of genes, which has been shown in other systems to be associated with signaling pathways controlling cell growth and proliferation. The Lbras cDNA complemented ras2 function in Saccharomyces cerevisiae and had the ability to transform mammalian cells. Expression of Lbras, present as a single copy in the genome, was dependent upon interaction with host roots. Northern analysis showed that expression was detectable in L. bicolor 48 h after interaction as well as in the established mycorrhizal tissue. Phylogenetic analysis with other Ras proteins showed that Lbras is related most closely to Aras of Aspergillus nidulans.

scholarly journals A Transcriptomic Atlas of the Ectomycorrhizal Fungus Laccaria bicolor

2021 ◽  
Vol 9 (12) ◽  
pp. 2612
Author(s):  
Joske Ruytinx ◽  
Shingo Miyauchi ◽  
Sebastian Hartmann-Wittulsky ◽  
Maíra de Freitas Pereira ◽  
Frédéric Guinet ◽  
...  
Keyword(s):  
Developmental Stages ◽  
Morphological Differentiation ◽  
Functional Differentiation ◽  
Ectomycorrhizal Fungus ◽  
Fine Tuning ◽  
Laccaria Bicolor ◽  
Transcriptional Responses ◽  
Wide Range ◽  
Transcriptional Changes ◽  
Ecm Fungi

Trees are able to colonize, establish and survive in a wide range of soils through associations with ectomycorrhizal (EcM) fungi. Proper functioning of EcM fungi implies the differentiation of structures within the fungal colony. A symbiotic structure is dedicated to nutrient exchange and the extramatricular mycelium explores soil for nutrients. Eventually, basidiocarps develop to assure last stages of sexual reproduction. The aim of this study is to understand how an EcM fungus uses its gene set to support functional differentiation and development of specialized morphological structures. We examined the transcriptomes of Laccaria bicolor under a series of experimental setups, including the growth with Populus tremula x alba at different developmental stages, basidiocarps and free-living mycelium, under various conditions of N, P and C supply. In particular, N supply induced global transcriptional changes, whereas responses to P supply seemed to be independent from it. Symbiosis development with poplar is characterized by transcriptional waves. Basidiocarp development shares transcriptional signatures with other basidiomycetes. Overlaps in transcriptional responses of L. bicolor hyphae to a host plant and N/C supply next to co-regulation of genes in basidiocarps and mature mycorrhiza were detected. Few genes are induced in a single condition only, but functional and morphological differentiation rather involves fine tuning of larger gene sets. Overall, this transcriptomic atlas builds a reference to study the function and stability of EcM symbiosis in distinct conditions using L. bicolor as a model and indicates both similarities and differences with other ectomycorrhizal fungi, allowing researchers to distinguish conserved processes such as basidiocarp development from nutrient homeostasis.

Evaluation of the nucleopolyhedrovirus of Anticarsia gemmatalis as a vector for gene therapy in mammals

2020 ◽  
Vol 20 ◽  
Author(s):  
Cintia N. Parsza ◽  
Diego L. Mengual Gómez ◽  
Jorge Alejandro Simonin ◽  
Mariano Nicolás Belaich ◽  
Pablo Daniel Ghiringhelli
Keyword(s):  
Gene Therapy ◽  
Mammalian Cells ◽  
Insect Cells ◽  
Autographa Californica ◽  
Anticarsia Gemmatalis ◽  
Agricultural Pests ◽  
Mammalian Cell Lines ◽  
Delivery Vector ◽  
Wide Range ◽  
Insect Pathogens

Background: Baculoviruses are insect pathogens with important biotechnological applications that transcend their use as biological controllers of agricultural pests. One species, Autographa californica multiple nucleopolhyedrovirus (AcMNPV) has been extensively exploited as a molecular platform to produce recombinant proteins and as a delivery vector for genes in mammals, because it can transduce a wide range of mammalian cells and tissues without replicating or producing progeny. Objective/Method: To investigate if the budded virions of Anticarsia gemmatalis multiple nucleopolhyedrovirus (AgMNPV) species has the same ability, the viral genome was modified by homologous recombination into susceptible insect cells to integrate reporter genes and then it was evaluated on mammalian cell lines in comparative form with respect to equivalent viruses derived from AcMNPV. Besides, the replicative capacity of AgMNPV´s virions in mammals was determined. Results: The experiments carried out showed that the recombinant variant of AgMNPV transduces and support the expression of delivered genes but not replicates in mammalian cells. Conclusion: Consequently, this insect pathogen is proposed as an alternative of non-infectious viruses in humans to explore new approaches in gene therapy and other applications based on the use of mammalian cells.

scholarly journals Individual Expression of Hepatitis A Virus 3C Protease Induces Ferroptosis in Human Cells In Vitro

2021 ◽  
Vol 22 (15) ◽  
pp. 7906
Author(s):  
Alexey A. Komissarov ◽  
Maria A. Karaseva ◽  
Marina P. Roschina ◽  
Andrey V. Shubin ◽  
Nataliya A. Lunina ◽  
...  
Keyword(s):  
Cell Death ◽  
Hepatitis A ◽  
Hepatitis A Virus ◽  
Proteolytic Enzymes ◽  
Morphological Changes ◽  
Human Cells ◽  
Mitochondrial Potential ◽  
3C Protease ◽  
Wide Range

Regulated cell death (RCD) is a fundamental process common to nearly all living beings and essential for the development and tissue homeostasis in animals and humans. A wide range of molecules can induce RCD, including a number of viral proteolytic enzymes. To date, numerous data indicate that picornaviral 3C proteases can induce RCD. In most reported cases, these proteases induce classical caspase-dependent apoptosis. In contrast, the human hepatitis A virus 3C protease (3Cpro) has recently been shown to cause caspase-independent cell death accompanied by previously undescribed features. Here, we expressed 3Cpro in HEK293, HeLa, and A549 human cell lines to characterize 3Cpro-induced cell death morphologically and biochemically using flow cytometry and fluorescence microscopy. We found that dead cells demonstrated necrosis-like morphological changes including permeabilization of the plasma membrane, loss of mitochondrial potential, as well as mitochondria and nuclei swelling. Additionally, we showed that 3Cpro-induced cell death was efficiently blocked by ferroptosis inhibitors and was accompanied by intense lipid peroxidation. Taken together, these results indicate that 3Cpro induces ferroptosis upon its individual expression in human cells. This is the first demonstration that a proteolytic enzyme can induce ferroptosis, the recently discovered and actively studied type of RCD.

scholarly journals An Evaluation of Morphological Changes and Deformability of Suspended Red Blood Cells Prepared Using Whole Blood with Different Hemoglobin Levels of Tibetans

2021 ◽  
pp. 1-10
Author(s):  
Rui Zhong ◽  
Dingding Han ◽  
Xiaodong Wu ◽  
Hong Wang ◽  
Wanjing Li ◽  
...  
Keyword(s):  
Red Blood Cells ◽  
Whole Blood ◽  
Blood Cells ◽  
Morphological Changes ◽  
Osmotic Fragility ◽  
Spherical Shape ◽  
Hypoxic Environment ◽  
Wide Range ◽  
Group A ◽  
Cell Membrane Protein

Background: The hypoxic environment stimulates the human body to increase the levels of hemoglobin (HGB) and hematocrit and the number of red blood cells. Such enhancements have individual differences, leading to a wide range of HGB in Tibetans’ whole blood (WB). Study Design: WB of male Tibetans was divided into 3 groups according to different HGB (i.e., A: >120 but ≤185 g/L, B: >185 but ≤210 g/L, and C: >210 g/L). Suspended red blood cells (SRBC) processed by collected WB and stored in standard conditions were examined aseptically on days 1, 14, 21, and 35 after storage. The routine biochemical indexes, deformability, cell morphology, and membrane proteins were tested. Results: Mean corpuscular volume, adenosine triphosphate, pH, and deformability were not different in group A vs. those in storage (p > 0.05). The increased rate of irreversible morphology of red blood cells was different among the 3 groups, but there was no difference in the percentage of red blood cells with an irreversible morphology after 35 days of storage. Group C performed better in terms of osmotic fragility and showed a lower rigid index than group A. Furthermore, SDS-PAGE revealed similar cross-linking degrees of cell membrane protein but the band 3 protein of group C seemed to experience weaker clustering than that of group A as detected by Western Blot analysis after 35 days of storage. Conclusions: There was no difference in deformability or morphological changes in the 3 groups over the 35 days of storage. High HGB levels of plateau SRBC did not accelerate the RBC change from a biconcave disc into a spherical shape and it did not cause a reduction in deformability during 35 days of preservation in bank conditions.

scholarly journals Autophagy Deficiency by Atg4B Loss Leads to Metabolomic Alterations in Mice

Metabolites ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 481
Author(s):  
Gemma G. Martínez-García ◽  
Raúl F. Pérez ◽  
Álvaro F. Fernández ◽  
Sylvere Durand ◽  
Guido Kroemer ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Amino Acids ◽  
Mammalian Cells ◽  
Tissue Homeostasis ◽  
In Vivo Studies ◽  
Protective Mechanism ◽  
Cardiac Damage ◽  
Wide Range ◽  
First Time

Autophagy is an essential protective mechanism that allows mammalian cells to cope with a variety of stressors and contributes to maintaining cellular and tissue homeostasis. Due to these crucial roles and also to the fact that autophagy malfunction has been described in a wide range of pathologies, an increasing number of in vivo studies involving animal models targeting autophagy genes have been developed. In mammals, total autophagy inactivation is lethal, and constitutive knockout models lacking effectors of this route are not viable, which has hindered so far the analysis of the consequences of a systemic autophagy decline. Here, we take advantage of atg4b−/− mice, an autophagy-deficient model with only partial disruption of the process, to assess the effects of systemic reduction of autophagy on the metabolome. We describe for the first time the metabolic footprint of systemic autophagy decline, showing that impaired autophagy results in highly tissue-dependent alterations that are more accentuated in the skeletal muscle and plasma. These changes, which include changes in the levels of amino-acids, lipids, or nucleosides, sometimes resemble those that are frequently described in conditions like aging, obesity, or cardiac damage. We also discuss different hypotheses on how impaired autophagy may affect the metabolism of several tissues in mammals.

scholarly journals Isolation and characterization of multiple variants of recombinant human interleukin 4 expressed in mammalian cells.

1988 ◽  
Vol 263 (22) ◽  
pp. 10817-10823
Author(s):  
H V Le ◽  
L Ramanathan ◽  
J E Labdon ◽  
C A Mays-Ichinco ◽  
R Syto ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Interleukin 4 ◽  
Isolation And Characterization ◽  
Multiple Variants

scholarly journals Isolation and Characterization of Strain Exiguobacterium sp. KRL4, a Producer of Bioactive Secondary Metabolites from a Tibetan Glacier

2021 ◽  
Vol 9 (5) ◽  
pp. 890
Author(s):  
Pietro Tedesco ◽  
Fortunato Palma Esposito ◽  
Antonio Masino ◽  
Giovanni Andrea Vitale ◽  
Emiliana Tortorella ◽  
...  
Keyword(s):  
Phenotypic Analysis ◽  
Biological Assays ◽  
C Elegans ◽  
Isolation And Characterization ◽  
Wide Range ◽  
Protein Encoding ◽  
Nematocidal Activity ◽  
Extremophilic Microorganisms ◽  
Unique Source

Extremophilic microorganisms represent a unique source of novel natural products. Among them, cold adapted bacteria and particularly alpine microorganisms are still underexplored. Here, we describe the isolation and characterization of a novel Gram-positive, aerobic rod-shaped alpine bacterium (KRL4), isolated from sediments from the Karuola glacier in Tibet, China. Complete phenotypic analysis was performed revealing the great adaptability of the strain to a wide range of temperatures (5–40 °C), pHs (5.5–8.5), and salinities (0–15% w/v NaCl). Genome sequencing identified KRL4 as a member of the placeholder genus Exiguobacterium_A and annotation revealed that only half of the protein-encoding genes (1522 of 3079) could be assigned a putative function. An analysis of the secondary metabolite clusters revealed the presence of two uncharacterized phytoene synthase containing pathways and a novel siderophore pathway. Biological assays confirmed that the strain produces molecules with antioxidant and siderophore activities. Furthermore, intracellular extracts showed nematocidal activity towards C. elegans, suggesting that strain KRL4 is a source of anthelmintic compounds.

scholarly journals A novel human Smad4 mutation is involved in papillary thyroid carcinoma progression

2011 ◽  
Vol 19 (1) ◽  
pp. 39-55 ◽  
Author(s):  
Sonia D'Inzeo ◽  
Arianna Nicolussi ◽  
Caterina Francesca Donini ◽  
Massimo Zani ◽  
Patrizia Mancini ◽  
...  
Keyword(s):  
Papillary Thyroid Carcinoma ◽  
Thyroid Carcinoma ◽  
Mammalian Cells ◽  
Transforming Growth Factor ◽  
Human Cancer ◽  
Papillary Thyroid ◽  
Tgfβ Signaling ◽  
Strong Reduction ◽  
Mesenchymal Transition ◽  
Wide Range

Smad proteins are the key effectors of the transforming growth factor β (TGFβ) signaling pathway in mammalian cells. Smad4 plays an important role in human physiology, and its mutations were found with high frequency in wide range of human cancer. In this study, we have functionally characterized Smad4 C324Y mutation, isolated from a nodal metastasis of papillary thyroid carcinoma. We demonstrated that the stable expression of Smad4 C324Y in FRTL-5 cells caused a significant activation of TGFβ signaling, responsible for the acquisition of transformed phenotype and invasive behavior. The coexpression of Smad4 C324Y with Smad4 wild-type determined an increase of homo-oligomerization of Smad4 with receptor-regulated Smads and a lengthening of nuclear localization. FRTL-5 clones overexpressing Smad4 C324Y showed a strong reduction of response to antiproliferative action of TGFβ1, acquired the ability to grow in anchorage-independent conditions, showed a fibroblast-like appearance and a strong reduction of the level of E-cadherin, one crucial event of the epithelial–mesenchymal transition process. The acquisition of a mesenchymal phenotype gave the characteristics of increased cellular motility and a significant reduction in adhesion to substrates such as fibronectin and laminin. Overall, our results demonstrate that the Smad4 C324Y mutation plays an important role in thyroid carcinogenesis and can be considered as a new prognostic and therapeutic target for thyroid cancer.

scholarly journals Isolation and Characterization of Bacteriophage ZCSE6 against Salmonella spp.: Phage Application in Milk

Biologics ◽  
2021 ◽  
Vol 1 (2) ◽  
pp. 164-176
Author(s):  
Abdallah S. Abdelsattar ◽  
Anan Safwat ◽  
Rana Nofal ◽  
Amera Elsayed ◽  
Salsabil Makky ◽  
...  
Keyword(s):  
Pathogenic Bacteria ◽  
Raw Milk ◽  
Salmonella Spp ◽  
Isolation And Characterization ◽  
Food Borne ◽  
Wide Range ◽  
Almost All ◽  
And Control ◽  
Milk And Dairy Products

Food safety is very important in the food industry as most pathogenic bacteria can cause food-borne diseases and negatively affect public health. In the milk industry, contamination with Salmonella has always been a challenge, but the risks have dramatically increased as almost all bacteria now show resistance to a wide range of commercial antibiotics. This study aimed to isolate a bacteriophage to be used as a bactericidal agent against Salmonella in milk and dairy products. Here, phage ZCSE6 has been isolated from raw milk sample sand molecularly and chemically characterized. At different multiplicities of infection (MOIs) of 0.1, 0.01, and 0.001, the phage–Salmonella interaction was studied for 6 h at 37 °C and 24 h at 8 °C. In addition, ZCSE6 was tested against Salmonella contamination in milk to examine its lytic activity for 3 h at 37 °C. The results showed that ZCSE6 has a small genome size (<48.5 kbp) and belongs to the Siphovirus family. Phage ZCSE6 revealed a high thermal and pH stability at various conditions that mimic milk manufacturing and supply chain conditions. It also demonstrated a significant reduction in Salmonella concentration in media at various MOIs, with higher bacterial eradication at higher MOI. Moreover, it significantly reduced Salmonella growth (MOI 1) in milk, manifesting a 1000-fold decrease in bacteria concentration following 3 h incubation at 37 °C. The results highlighted the strong ability of ZCSE6 to kill Salmonella and control its growth in milk. Thus, ZCSE6 is recommended as a biocontrol agent in milk to limit bacterial growth and increase the milk shelf-life.

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