scholarly journals Mechanism of the Affinity-Enhancing Effect of Isatin on Human Ferrochelatase and Adrenodoxin Reductase Complex Formation: Implication for Protein Interactome Regulation

2020 ◽  
Vol 21 (20) ◽  
pp. 7605
Author(s):  
Pavel V. Ershov ◽  
Alexander V. Veselovsky ◽  
Yuri V. Mezentsev ◽  
Evgeniy O. Yablokov ◽  
Leonid A. Kaluzhskiy ◽  
...  
Keyword(s):  
Complex Formation ◽  
Protein Interactions ◽  
Wide Spectrum ◽  
Three Dimensional ◽  
3D Models ◽  
Bioinformatic Analysis ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Interacting Protein ◽  
Enhancing Effect

Isatin (indole-2, 3-dione) is a non-peptide endogenous bioregulator exhibiting a wide spectrum of biological activity, realized in the cell via interactions with numerous isatin-binding proteins, their complexes, and (sub) interactomes. There is increasing evidence that isatin may be involved in the regulation of complex formations by modulating the affinity of the interacting protein partners. Recently, using Surface Plasmon Resonance (SPR) analysis, we have found that isatin in a concentration dependent manner increased interaction between two human mitochondrial proteins, ferrochelatase (FECH), and adrenodoxine reductase (ADR). In this study, we have investigated the affinity-enhancing effect of isatin on the FECH/ADR interaction. The SPR analysis has shown that FECH forms not only homodimers, but also FECH/ADR heterodimers. The affinity-enhancing effect of isatin on the FECH/ADR interaction was highly specific and was not reproduced by structural analogues of isatin. Bioinformatic analysis performed using three dimensional (3D) models of the interacting proteins and in silico molecular docking revealed the most probable mechanism involving FECH/isatin/ADR ternary complex formation. In this complex, isatin is targeted to the interface of interacting FECH and ADR monomers, forming hydrogen bonds with both FECH and ADR. This is a new regulatory mechanism by which isatin can modulate protein–protein interactions (PPI).

scholarly journals Modulatory Effects of Osthole on Lipopolysaccharides-Induced Inflammation in Caco-2 Cell Monolayer and Co-Cultures with THP-1 and THP-1-Derived Macrophages

Nutrients ◽  
2020 ◽  
Vol 13 (1) ◽  
pp. 123
Author(s):  
Natalia K. Kordulewska ◽  
Justyna Topa ◽  
Małgorzata Tańska ◽  
Anna Cieślińska ◽  
Ewa Fiedorowicz ◽  
...  
Keyword(s):  
Gene Expression ◽  
Inflammatory Reaction ◽  
Wide Spectrum ◽  
Cell Monolayer ◽  
In Vivo Studies ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Tnf Α

Lipopolysaccharydes (LPS) are responsible for the intestinal inflammatory reaction, as they may disrupt tight junctions and induce cytokines (CKs) secretion. Osthole has a wide spectrum of pharmacological effects, thus its anti-inflammatory potential in the LPS-treated Caco-2 cell line as well as in Caco-2/THP-1 and Caco-2/macrophages co-cultures was investigated. In brief, Caco-2 cells and co-cultures were incubated with LPS to induce an inflammatory reaction, after which osthole (150–450 ng/mL) was applied to reduce this effect. After 24 h, the level of secreted CKs and changes in gene expression were examined. LPS significantly increased the levels of IL-1β, -6, -8, and TNF-α, while osthole reduced this effect in a concentration-dependent manner, with the most significant decrease when a 450 ng/mL dose was applied (p < 0.0001). A similar trend was observed in changes in gene expression, with the significant osthole efficiency at a concentration of 450 ng/μL for IL1R1 and COX-2 (p < 0.01) and 300 ng/μL for NF-κB (p < 0.001). Osthole increased Caco-2 monolayer permeability, thus if it would ever be considered as a potential drug for minimizing intestinal inflammatory symptoms, its safety should be confirmed in extended in vitro and in vivo studies.

Direct interaction of the tau 1 transactivation domain of the human glucocorticoid receptor with the basal transcriptional machinery

1993 ◽  
Vol 13 (1) ◽  
pp. 399-407
Author(s):  
I J McEwan ◽  
A P Wright ◽  
K Dahlman-Wright ◽  
J Carlstedt-Duke ◽  
J A Gustafsson
Keyword(s):  
Glucocorticoid Receptor ◽  
Protein Interactions ◽  
Core Promoter ◽  
Specific Interaction ◽  
Direct Interaction ◽  
Dependent Manner ◽  
Transactivation Domain ◽  
Yeast Saccharomyces Cerevisiae ◽  
Concentration Dependent Manner

We have used a yeast (Saccharomyces cerevisiae) cell free transcription system to study protein-protein interactions involving the tau 1 transactivation domain of the human glucocorticoid receptor that are important for transcriptional transactivation by the receptor. Purified tau 1 specifically inhibited transcription from a basal promoter derived from the CYC1 gene and from the adenovirus 2 major late core promoter in a concentration-dependent manner. This inhibition or squelching was correlated with the transactivation activity of tau 1. Recombinant yeast TATA-binding protein (yTFIID), although active in vitro, did not specifically reverse the inhibitory effect of tau 1. In addition, no specific interaction between tau 1 and yTFIID could be shown in vitro by affinity chromatography. Taken together, these results indicate that the tau 1 transactivation domain of the human glucocorticoid receptor interacts directly with the general transcriptional apparatus through some target protein(s) that is distinct from the TATA-binding factor. Furthermore, this assay can be used to identify interacting factors, since after phosphocellulose chromatography of a whole-cell yeast extract, a fraction that contained an activity which selectively counteracted the squelching effect of tau 1 was found.

scholarly journals Tat-Cannabinoid Receptor Interacting Protein Reduces Ischemia-Induced Neuronal Damage and Its Possible Relationship with 14-3-3η

Cells ◽  
2020 ◽  
Vol 9 (8) ◽  
pp. 1827 ◽  
Author(s):  
Hyun Jung Kwon ◽  
Duk-Soo Kim ◽  
Woosuk Kim ◽  
Hyo Young Jung ◽  
Yeon Hee Yu ◽  
...  
Keyword(s):  
Cannabinoid Receptor ◽  
Neuronal Damage ◽  
Cell Damage ◽  
Ischemia Reperfusion ◽  
Ischemic Damage ◽  
Dependent Manner ◽  
Ht22 Cells ◽  
Concentration Dependent Manner ◽  
H2o2 Treatment ◽  
Interacting Protein

Cannabinoid receptor-interacting protein 1a (CRIP1a) binds to the C-terminal domain of cannabinoid 1 receptor (CB1R) and regulates CB1R activities. In this study, we made Tat-CRIP1a fusion proteins to enhance CRIP1a penetration into neurons and brain and to evaluate the function of CRIP1a in neuroprotection following oxidative stress in HT22 hippocampal cells and transient forebrain ischemia in gerbils. Purified exogenous Tat-CRIP1a was penetrated into HT22 cells in a time and concentration-dependent manner and prevented H2O2-induced reactive oxygen species formation, DNA fragmentation, and cell damage. Tat-CRIP1a fusion protein also ameliorated the reduction of 14-3-3η expression by H2O2 treatment in HT22 cells. Ischemia–reperfusion damage caused motor hyperactivity in the open field test of gerbils; however, the treatment of Tat-CRIP1a significantly reduced hyperactivity 1 day after ischemia. Four days after ischemia, the administration of Tat-CRIP1a restored the loss of pyramidal neurons and decreased reactive astrocytosis and microgliosis induced by ischemic damage in the hippocampal cornu Ammonis (CA)1 region. Ischemic damage decreased 14-3-3η expression in all hippocampal sub-regions 4 days after ischemia; however, the treatment of Tat-CRIP1 ameliorated the reduction of 14-3-3η expression. These results suggest that Tat-CRIP1a attenuates neuronal damage and hyperactivity induced by ischemic damage, and it restores normal expression levels of 14-3-3η protein in the hippocampus.

Nuclear receptor action involved with sex differentiation

2003 ◽  
Vol 75 (11-12) ◽  
pp. 1771-1784 ◽  
Author(s):  
I. A. Hughes ◽  
Howard Martin ◽  
Jarmo Jääskeläinen ◽  
C. L. Acerini
Keyword(s):  
Protein Interactions ◽  
Molecular Mechanisms ◽  
Reproductive Tract ◽  
Sex Differentiation ◽  
Disorders Of Sex Development ◽  
Nuclear Hormone Receptor ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Sex Development ◽  
Sequence Of Events

Sex determination and differentiation in the male is an orderly sequence of events coordinated by genetic and hormonal factors operating in a time- and concentration-dependent manner. The constitutive sex in mammals is female. Disorders of fetal sex development have provided the means to identify testis-determining genes and the molecular mechanisms of hormone action. Thus, the androgen receptor, a nuclear hormone receptor critical for androgen-induced male sex differentiation, displays unique intra-receptor and protein-protein interactions which, when disturbed, can result in extreme forms of sex reversal. Polymorphic variants are associated with milder disorders of sex development. Against this genetic background, endocrine active substances may further contribute to the underlying causes of an increase in male reproductive tract disorders.

scholarly journals Modulation of Calmodulin and Protein Kinase C Activities by Pencillium Mycotoxins

1999 ◽  
Vol 18 (2) ◽  
pp. 91-96 ◽  
Author(s):  
I. Pala ◽  
A. Srinivasan ◽  
P. J. S. Vig ◽  
D. Desaiah
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Complex Formation ◽  
Mammalian Brain ◽  
Dependent Manner ◽  
Active Conformation ◽  
Cellular Functions ◽  
Concentration Dependent Manner ◽  
Kinase C ◽  
Ic50 Values

Calmodulin (CaM), a calcium-binding protein, is found in high concentrations in mammalian brain where it plays a pivotal role in a large number of cellular functions. Protein kinase C (PKC), a multifunctional cytosolic enzyme, in the presence of both Ca2+ and phospholipids, transduce extracellular signals into intracellu-lar events. Both CaM and PKC are partially involved in maintaining Ca2+ homeostasis in the cell. Any fluctuations in the intracel-lular Ca2+ can modulate cellular functions and may contribute to neuronal dysfunction. Hence, the present investigation was initiated to study the effects of some selected penicillium (naturally occurring tremorgenic) mycotoxins like secalonic acid, citreoviridin, and verruculogen on CaM activity, active conformation of CaM and PKC activity. Stimulation of CaM-deflcient bovine brain 3′-5′ phosphodieste rase (PDE) indicated CaM activity. The modification of CaM active conformation was studied by the binding of fluorescent probe N-phenyl-1-napthylamine (NPN) to CaM. Alterations in the fluorescence of dansyl-CaM was used to study the effect of these compounds on complex formation between CaM and PDE. Rat brain cytosolic PKC was studied using 32P-ATP as a measure of altered protein phosphorylation. The concentrations of mycotoxins used were in the range of 10 to 50 μM. All three mycotoxins inhibited CaM-stimulated PDE activity in a concentration-dependent manner. Citreoviridin and secalonic acid inhibited NPN fluorescence and Ca2+-dependent complex formation of dansyl-CaM and PDE. The IC50 values for NPN fluorescence of citreoviridin and secalonic acid were 13 μM and 19 μM respectively. However, verruculogen showed little effect on NPN fluorescence and the Ca2+-dependent complex formation of dansyl-CaM and PDE. These mycotoxins also inhibited PKC activity in a concentration-dependent manner with IC50 values of 19.8, 25.7, and 38.4 μM for secalonic acid, citreoviridin, and verruculogen, respectively. The results of our study suggest that these mycotoxins at very low concentrations are interacting with CaM and PKC. Such an effect could lead to impairment of neurotransmission and result in neurotoxicity.

scholarly journals Organotypic Epithelial Raft Cultures as a Model for Evaluating Compounds against Alphaherpesviruses

2005 ◽  
Vol 49 (11) ◽  
pp. 4671-4680 ◽  
Author(s):  
Graciela Andrei ◽  
Joost van den Oord ◽  
Pierre Fiten ◽  
Ghislain Opdenakker ◽  
Chris De Wolf-Peeters ◽  
...  
Keyword(s):  
Antiviral Agents ◽  
Plaque Assay ◽  
Three Dimensional ◽  
Liquid Interface ◽  
Dependent Manner ◽  
Wild Type ◽  
Organotypic Cultures ◽  
Concentration Dependent Manner ◽  
Raft Culture ◽  
Air Liquid Interface

ABSTRACT The course of herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) and varicella-zoster virus (VZV) infections in squamous epithelial cells cultured in a three-dimensional organotypic raft culture was tested. In these raft cultures, normal human keratinocytes isolated from neonatal foreskins grown at the air-liquid interface stratified and differentiated, reproducing a fully differentiated epithelium. Typical cytopathic changes identical to those found in the squamous epithelium in vivo, including ballooning and reticular degeneration with the formation of multinucleate cells, were observed throughout the raft following infection with HSV and VZV at different times after lifting the cultures to the air-liquid interface. For VZV, the aspects of the lesions depended on the stage of differentiation of the organotypic cultures. The activity of reference antiviral agents, acyclovir (ACV), penciclovir (PCV), brivudin (BVDU), foscarnet (PFA), and cidofovir (CDV), was evaluated against wild-type and thymidine kinase (TK) mutants of HSV and VZV in the raft cultures. ACV, PCV, and BVDU protected the epithelium against cytopathic effect induced by wild-type viruses in a concentration-dependent manner, while treatment with CDV and PFA proved protective against the cytodestructive effects induced by both TK+ and TK− strains. The quantification of the antiviral effects in the rafts were accomplished by measuring viral titers by plaque assay for HSV and by measuring viral DNA load by real-time PCR for VZV. A correlation between the degree of protection as determined by histological examination and viral quantification could be demonstrated The three-dimensional epithelial raft culture represents a novel model for the study of antiviral agents active against HSV and VZV. Since no animal model is available for the evaluation of antiviral agents against VZV, the organotypic cultures may be considered a model to evaluate the efficacy of new anti-VZV antivirals before clinical trials.

scholarly journals Control of Formation and Cellular Detachment from Shewanella oneidensis MR-1 Biofilms by Cyclic di-GMP

2006 ◽  
Vol 188 (7) ◽  
pp. 2681-2691 ◽  
Author(s):  
Kai M. Thormann ◽  
Stefanie Duttler ◽  
Renee M. Saville ◽  
Mamoru Hyodo ◽  
Soni Shukla ◽  
...  
Keyword(s):  
Ectopic Expression ◽  
Three Dimensional ◽  
Shewanella Oneidensis ◽  
Dependent Manner ◽  
Gene Encoding ◽  
Concentration Dependent Manner ◽  
Environmental Perturbations ◽  
Environmental Trigger ◽  
Exclusive Processes ◽  
Stability And Resilience

ABSTRACT Stability and resilience against environmental perturbations are critical properties of medical and environmental biofilms and pose important targets for their control. Biofilm stability is determined by two mutually exclusive processes: attachment of cells to and detachment from the biofilm matrix. Using Shewanella oneidensis MR-1, an environmentally versatile, Fe(III) and Mn(IV) mineral-reducing microorganism, we identified mxdABCD as a new set of genes essential for formation of a three-dimensional biofilm. Molecular analysis revealed that mxdA encodes a cyclic bis(3′,5′)guanylic acid (cyclic di-GMP)-forming enzyme with an unusual GGDEF motif, i.e., NVDEF, which is essential for its function. mxdB encodes a putative membrane-associated glycosyl transferase. Both genes are essential for matrix attachment. The attachment-deficient phenotype of a ΔmxdA mutant was rescued by ectopic expression of VCA0956, encoding another diguanylate cyclase. Interestingly, a rapid cellular detachment from the biofilm occurred upon induction of yhjH, a gene encoding an enzyme that has been shown to have phosphodiesterase activity. In this way, it was possible to bypass the previously identified sudden depletion of molecular oxygen as an environmental trigger to induce biofilm dissolution. We propose a model for c-di-GMP as a key intracellular regulator for controlling biofilm stability by shifting the state of a biofilm cell between attachment and detachment in a concentration-dependent manner.

Chloroplast division protein ARC3 acts on FtsZ2 by preventing filament bundling and enhancing GTPase activity

2018 ◽  
Vol 475 (1) ◽  
pp. 99-115 ◽  
Author(s):  
Rahamthulla S. Shaik ◽  
Min Woo Sung ◽  
Stanislav Vitha ◽  
Andreas Holzenburg
Keyword(s):  
Sequence Similarity ◽  
Three Dimensional ◽  
Chloroplast Division ◽  
Particle Analysis ◽  
Temperature Sensitive ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Z Ring

Chloroplasts evolved from cyanobacterial endosymbiotic ancestors and their division is a complex process initiated by the assembly of cytoskeletal FtsZ (Filamentous temperature sensitive Z) proteins into a ring structure at the division site (Z-ring). The cyanobacterial Z-ring positioning system (MinCDE proteins) is also conserved in chloroplasts, except that MinC was lost and replaced by the eukaryotic ARC3 (accumulation and replication of chloroplasts). Both MinC and ARC3 act as negative regulators of FtsZ assembly, but ARC3 bears little sequence similarity with MinC. Here, light scattering assays, co-sedimentation, GTPase assay and transmission electron microscopy in conjunction with single-particle analysis have been used to elucidate the structure of ARC3 and its effect on its main target in chloroplast division, FtsZ2. Analysis of FtsZ2 in vitro assembly reactions in the presence and absence of GMPCPP showed that ARC3 promotes FtsZ2 debundling and disassembly of existing filaments in a concentration-dependent manner and requires GTP hydrolysis. Three-dimensional reconstruction of ARC3 revealed an almost circular molecule in which the FtsZ-binding N-terminus and the C-terminal PARC6 (paralog of ARC6)-binding MORN (Membrane Occupation and Recognition Nexus) domain are in close proximity and suggest a model for PARC6-enabled binding of ARC3 to FtsZ2. The latter is corroborated by in vivo data.

scholarly journals Proton-Pumping-ATPase-Targeted Antifungal Activity of a Novel Conjugated Styryl Ketone

1999 ◽  
Vol 43 (12) ◽  
pp. 2950-2959 ◽  
Author(s):  
Elias K. Manavathu ◽  
Jonathan R. Dimmock ◽  
Sarvesh C. Vashishtha ◽  
Pranatharthi H. Chandrasekar
Keyword(s):  
Antifungal Activity ◽  
External Medium ◽  
Wide Spectrum ◽  
Pathogenic Fungi ◽  
Candida Guilliermondii ◽  
Proton Pumping ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Membrane Components ◽  
Hydrolysis Of

ABSTRACT NC1175 (3-[3-(4-chlorophenyl)-2-propenoyl]-4-[2-(4-chlorophenyl)vinylene]-1-ethyl-4-piperidinol hydrochloride) is a novel thiol-blocking conjugated styryl ketone that exhibits activity against a wide spectrum of pathogenic fungi. Incubation of NC1175 with various concentrations of cysteine and glutathione eliminated its antifungal activity in a concentration-dependent fashion. Since NC1175 is a lipophilic compound that has the potential to interact with cytoplasmic membrane components, we examined its effect on the membrane-located proton-translocating ATPase (H+-ATPase) of yeast (Candida albicans, Candida krusei,Candida guilliermondii, Candida glabrata, andSaccharomyces cerevisiae) and Aspergillus(Aspergillus fumigatus, Aspergillus niger,Aspergillus flavus, and Aspergillus nidulans) species. The glucose-induced acidification of external medium due to H+-ATPase-mediated expulsion of intracellular protons by these fungi was measured in the presence of several concentrations of the drug. NC1175 (12.5 to 50 μM) inhibited acidification of external medium by Candida, Saccharomyces, andAspergillus species in a concentration-dependent manner. Vanadate-inhibited hydrolysis of ATP by membrane fractions of C. albicans was completely inhibited by 50 μM NC1175, suggesting that the target of action of NC1175 in these fungi may include H+-ATPase.

scholarly journals Effect of the divalent cations zinc and calcium on the structure and mechanics of reconstituted vimentin intermediate filaments

2019 ◽  
Author(s):  
Huayin Wu ◽  
Yinan Shen ◽  
Dianzhuo Wang ◽  
Harald Herrmann ◽  
Robert D. Goldman ◽  
...  
Keyword(s):  
Intermediate Filaments ◽  
Network Structure ◽  
Protein Interactions ◽  
Divalent Cations ◽  
Dependent Manner ◽  
Structural Elements ◽  
Concentration Dependent Manner ◽  
Hofmeister Effect ◽  
Ion Species

AbstractDivalent cations in a concentration-dependent manner behave as effective crosslinkers of intermediate filaments (IFs) such as vimentin IF (VIF). These interactions have been mostly attributed to their multivalency. However, ion-protein interactions often depend on the ion species, and these effects have not been widely studied in IFs. Here we investigate the effects of two biologically important divalent cations, Zn2+ and Ca2+, on VIF network structure and mechanics in vitro. We find that the network structure is unperturbed at micromolar Zn2+ concentrations, but strong bundle formation is observed at a concentration of 100 μM. Microrheological measurements show that network stiffness increases with cation concentration. However, bundling of filaments softens the network. This trend also holds for VIF networks formed in the presence of Ca2+, but remarkably, a concentration of Ca2+ that is two orders higher is needed to achieve the same effect as with Zn2+, which suggests the importance of salt-protein interactions as described by the Hofmeister effect. Furthermore, we find evidence of competitive binding between the two divalent ion species. Hence, specific interactions between VIFs and divalent cations are likely to be an important mechanism by which cells can control their cytoplasmic mechanics.SignificanceIntermediate filaments are key structural elements within cells; they are known to form networks that can be crosslinked by divalent cations, but the interactions between the ions and the filaments are not well understood. By measuring the effects that two divalent cations, zinc and calcium, have on the structure and mechanics of vimentin intermediate filaments (VIFs), we show that although both have concentration-dependent effects on VIFs, much more calcium is needed to achieve the same effect as a small amount of zinc. Furthermore, when mixtures of the ions are present, the results suggest that there is binding competition. Thus, cells may use the presence of different cation species to precisely control their internal mechanical properties.

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