scholarly journals Dictyostelium Differentiation-Inducing Factor-1 Promotes Glucose Uptake, at Least in Part, via an AMPK-Dependent Pathway in Mouse 3T3-L1 Cells

2021 ◽  
Vol 22 (5) ◽  
pp. 2293
Author(s):  
Yuzuru Kubohara ◽  
Yoshimi Homma ◽  
Hiroshi Shibata ◽  
Yoshiteru Oshima ◽  
Haruhisa Kikuchi
Keyword(s):  
Glucose Uptake ◽  
Mammalian Cells ◽  
Glucose Consumption ◽  
Cellular Slime Mold ◽  
Dependent Manner ◽  
Metabolome Analysis ◽  
Compound C ◽  
Dependent Pathway

Differentiation-inducing factor-1 (DIF-1) is a chlorinated alkylphenone (a polyketide) found in the cellular slime mold Dictyostelium discoideum. DIF-1 and its derivative, DIF-1(3M) promote glucose consumption in vitro in mammalian cells and in vivo in diabetic rats; they are expected to be the leading antiobesity and antidiabetes compounds. In this study, we investigated the mechanisms underlying the actions of DIF-1 and DIF-1(3M). In isolated mouse liver mitochondria, these compounds at 2–20 mM promoted oxygen consumption in a dose-dependent manner, suggesting that they act as mitochondrial uncouplers, whereas CP-DIF-1 (another derivative of DIF-1) at 10–20 mM had no effect. In confluent mouse 3T3-L1 fibroblasts, DIF-1 and DIF-1(3M) but not CP-DIF-1 induced phosphorylation (and therefore activation) of AMP kinase (AMPK) and promoted glucose consumption and metabolism. The DIF-induced glucose consumption was reduced by compound C (an AMPK inhibitor) or AMPK knock down. These data suggest that DIF-1 and DIF-1(3M) promote glucose uptake, at least in part, via an AMPK-dependent pathway in 3T3-L1 cells, whereas cellular metabolome analysis revealed that DIF-1 and DIF-1(3M) may act differently at least in part.

scholarly journals CP-31398 Restores DNA-binding Activity to Mutant p53in Vitrobut Does Not Affect p53 Homologs p63 and p73

2004 ◽  
Vol 279 (44) ◽  
pp. 45887-45896 ◽  
Author(s):  
Mark J. Demma ◽  
Serena Wong ◽  
Eugene Maxwell ◽  
Bimalendu Dasmahapatra
Keyword(s):  
Dna Binding ◽  
Mammalian Cells ◽  
P53 Protein ◽  
Binding Activity ◽  
Mutant P53 ◽  
Dependent Manner ◽  
Wild Type ◽  
Dna Binding Activity

The p53 protein plays a major role in the maintenance of genome stability in mammalian cells. Mutations of p53 occur in over 50% of all cancers and are indicative of highly aggressive cancers that are hard to treat. Recently, there has been a high degree of interest in therapeutic approaches to restore growth suppression functions to mutant p53. Several compounds have been reported to restore wild type function to mutant p53. One such compound, CP-31398, has been shown effectivein vivo, but questions have arisen to whether it actually affects p53. Here we show that mutant p53, isolated from cells treated with CP-31398, is capable of binding to p53 response elementsin vitro. We also show the compound restores DNA-binding activity to mutant p53 in cells as determined by a chromatin immunoprecipitation assay. In addition, using purified p53 core domain from two different hotspot mutants (R273H and R249S), we show that CP-31398 can restore DNA-binding activity in a dose-dependent manner. Using a quantitative DNA binding assay, we also show that CP-31398 increases significantly the amount of mutant p53 that binds to cognate DNA (Bmax) and its affinity (Kd) for DNA. The compound, however, does not affect the affinity (Kdvalue) of wild type p53 for DNA and only increasesBmaxslightly. In a similar assay PRIMA1 does not have any effect on p53 core DNA-binding activity. We also show that CP-31398 had no effect on the DNA-binding activity of p53 homologs p63 and p73.

scholarly journals SMG-1 Is a Phosphatidylinositol Kinase-Related Protein Kinase Required for Nonsense-Mediated mRNA Decay in Caenorhabditis elegans

2004 ◽  
Vol 24 (17) ◽  
pp. 7483-7490 ◽  
Author(s):  
Andrew Grimson ◽  
Sean O'Connor ◽  
Carrie Loushin Newman ◽  
Philip Anderson
Keyword(s):  
Protein Kinase ◽  
Mammalian Cells ◽  
Mrna Decay ◽  
Null Alleles ◽  
Dependent Manner ◽  
Messenger Rnas ◽  
Phosphatidylinositol Kinase ◽  
Nonsense Mediated Mrna Decay

ABSTRACT Eukaryotic messenger RNAs containing premature stop codons are selectively and rapidly degraded, a phenomenon termed nonsense-mediated mRNA decay (NMD). Previous studies with both Caenohabditis elegans and mammalian cells indicate that SMG-2/human UPF1, a central regulator of NMD, is phosphorylated in an SMG-1-dependent manner. We report here that smg-1, which is required for NMD in C. elegans, encodes a protein kinase of the phosphatidylinositol kinase superfamily of protein kinases. We identify null alleles of smg-1 and demonstrate that SMG-1 kinase activity is required in vivo for NMD and in vitro for SMG-2 phosphorylation. SMG-1 and SMG-2 coimmunoprecipitate from crude extracts, and this interaction is maintained in smg-3 and smg-4 mutants, both of which are required for SMG-2 phosphorylation in vivo and in vitro. SMG-2 is located diffusely through the cytoplasm, and its location is unaltered in mutants that disrupt the cycle of SMG-2 phosphorylation. We discuss the role of SMG-2 phosphorylation in NMD.

Inhibition of Hemoglobin Degrading Protease Falcipain-2 as a Mechanism for Anti-Malarial Activity of Triazole-Amino Acid Hybrids

2020 ◽  
Vol 20 (5) ◽  
pp. 377-389 ◽  
Author(s):  
Vigyasa Singh ◽  
Rahul Singh Hada ◽  
Amad Uddin ◽  
Babita Aneja ◽  
Mohammad Abid ◽  
...  
Keyword(s):  
Amino Acid ◽  
Cysteine Protease ◽  
Mammalian Cells ◽  
Antimalarial Activity ◽  
Docking Studies ◽  
Dependent Manner ◽  
Trophozoite Stage ◽  
Hemoglobin Degradation

Background: Novel drug development against malaria parasite over old conventional antimalarial drugs is essential due to rapid and indiscriminate use of drugs, which led to the emergence of resistant strains. Methods: In this study, previously reported triazole-amino acid hybrids (13-18) are explored against Plasmodium falciparum as antimalarial agents. Among six compounds, 15 and 18 exhibited antimalarial activity against P. falciparum with insignificant hemolytic activity and cytotoxicity towards HepG2 mammalian cells. In molecular docking studies, both compounds bind into the active site of PfFP-2 and block its accessibility to the substrate that leads to the inhibition of target protein further supported by in vitro analysis. Results: Antimalarial half-maximal inhibitory concentration (IC50) of 15 and 18 compounds were found to be 9.26 μM and 20.62 μM, respectively. Blood stage specific studies showed that compounds, 15 and 18 are effective at late trophozoite stage and block egress pathway of parasites. Decreased level of free monomeric heme was found in a dose dependent manner after the treatment with compounds 15 and 18, which was further evidenced by the reduction in percent of hemoglobin hydrolysis. Compounds 15 and 18 hindered hemoglobin degradation via intra- and extracellular cysteine protease falcipain-2 (PfFP-2) inhibitory activity both in in vitro and in vivo in P. falciparum. Conclusion: We report antimalarial potential of triazole-amino acid hybrids and their role in the inhibition of cysteine protease PfFP-2 as its mechanistic aspect.

scholarly journals Recruitment of the SWI-SNF Chromatin Remodeling Complex as a Mechanism of Gene Activation by the Glucocorticoid Receptor τ1 Activation Domain

2000 ◽  
Vol 20 (6) ◽  
pp. 2004-2013 ◽  
Author(s):  
Annika E. Wallberg ◽  
Kristen E. Neely ◽  
Ahmed H. Hassan ◽  
Jan-Åke Gustafsson ◽  
Jerry L. Workman ◽  
...  
Keyword(s):  
Glucocorticoid Receptor ◽  
Mammalian Cells ◽  
Gene Activation ◽  
Dependent Manner ◽  
Transactivation Domain ◽  
Chromatin Remodeling Complex ◽  
Nucleosomal Dna ◽  
Target Promoters

ABSTRACT The SWI-SNF complex has been shown to alter nucleosome conformation in an ATP-dependent manner, leading to increased accessibility of nucleosomal DNA to transcription factors. In this study, we show that the SWI-SNF complex can potentiate the activity of the glucocorticoid receptor (GR) through the N-terminal transactivation domain, τ1, in both yeast and mammalian cells. GR-τ1 can directly interact with purified SWI-SNF complex, and mutations in τ1 that affect the transactivation activity in vivo also directly affect τ1 interaction with SWI-SNF. Furthermore, the SWI-SNF complex can stimulate τ1-driven transcription from chromatin templates in vitro. Taken together, these results support a model in which the GR can directly recruit the SWI-SNF complex to target promoters during glucocorticoid-dependent gene activation. We also provide evidence that the SWI-SNF and SAGA complexes represent independent pathways of τ1-mediated activation but play overlapping roles that are able to compensate for one another under some conditions.

scholarly journals AMPylation matches BiP activity to client protein load in the endoplasmic reticulum

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Steffen Preissler ◽  
Cláudia Rato ◽  
Ruming Chen ◽  
Robin Antrobus ◽  
Shujing Ding ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Mammalian Cells ◽  
Atp Hydrolysis ◽  
Covalent Modification ◽  
Dependent Manner ◽  
Gene Encoding ◽  
Unfolded Protein ◽  
Peptide Dissociation

The endoplasmic reticulum (ER)-localized Hsp70 chaperone BiP affects protein folding homeostasis and the response to ER stress. Reversible inactivating covalent modification of BiP is believed to contribute to the balance between chaperones and unfolded ER proteins, but the nature of this modification has so far been hinted at indirectly. We report that deletion of FICD, a gene encoding an ER-localized AMPylating enzyme, abolished detectable modification of endogenous BiP enhancing ER buffering of unfolded protein stress in mammalian cells, whilst deregulated FICD activity had the opposite effect. In vitro, FICD AMPylated BiP to completion on a single residue, Thr518. AMPylation increased, in a strictly FICD-dependent manner, as the flux of proteins entering the ER was attenuated in vivo. In vitro, Thr518 AMPylation enhanced peptide dissociation from BiP 6-fold and abolished stimulation of ATP hydrolysis by J-domain cofactor. These findings expose the molecular basis for covalent inactivation of BiP.

scholarly journals Dual Regulation of c-Myc by p300 via Acetylation-Dependent Control of Myc Protein Turnover and Coactivation of Myc-Induced Transcription

2005 ◽  
Vol 25 (23) ◽  
pp. 10220-10234 ◽  
Author(s):  
Francesco Faiola ◽  
Xiaohui Liu ◽  
Szuying Lo ◽  
Songqin Pan ◽  
Kangling Zhang ◽  
...  
Keyword(s):  
Dna Binding ◽  
Cell Fate ◽  
Mammalian Cells ◽  
Dependent Manner ◽  
Dual Roles ◽  
Human Telomerase Reverse Transcriptase ◽  
Lysine Residues ◽  
Human Telomerase

ABSTRACT The c-Myc oncoprotein (Myc) controls cell fate by regulating gene transcription in association with a DNA-binding partner, Max. While Max lacks a transcription regulatory domain, the N terminus of Myc contains a transcription activation domain (TAD) that recruits cofactor complexes containing the histone acetyltransferases (HATs) GCN5 and Tip60. Here, we report a novel functional interaction between Myc TAD and the p300 coactivator-acetyltransferase. We show that p300 associates with Myc in mammalian cells and in vitro through direct interactions with Myc TAD residues 1 to 110 and acetylates Myc in a TAD-dependent manner in vivo at several lysine residues located between the TAD and DNA-binding domain. Moreover, the Myc:Max complex is differentially acetylated by p300 and GCN5 and is not acetylated by Tip60 in vitro, suggesting distinct functions for these acetyltransferases. Whereas p300 and CBP can stabilize Myc independently of acetylation, p300-mediated acetylation results in increased Myc turnover. In addition, p300 functions as a coactivator that is recruited by Myc to the promoter of the human telomerase reverse transcriptase gene, and p300/CBP stimulates Myc TAD-dependent transcription in a HAT domain-dependent manner. Our results suggest dual roles for p300/CBP in Myc regulation: as a Myc coactivator that stabilizes Myc and as an inducer of Myc instability via direct Myc acetylation.

scholarly journals In vitro antidiabetic, anti-inflammatory and antioxidant potential of the ethanol extract of Uromastyx hardwickii skin

2021 ◽  
Vol 18 (10) ◽  
pp. 2109-2115
Author(s):  
Waqas Ahmad Shams ◽  
Gauhar Rehman ◽  
Samuel Okwudili Onoja ◽  
Abid Ali ◽  
Khurshaid Khan ◽  
...  
Keyword(s):  
Hydrogen Peroxide ◽  
Glucose Uptake ◽  
Ethanol Extract ◽  
Antioxidant Potential ◽  
Yeast Cells ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Anti Inflammatory

Purpose: To evaluate the in vitro antidiabetic, anti-inflammatory and antioxidant potential of the ethanol extract of Uromastyx hardwickii Skin (UHSEE). Methods: The in vitro effects of UHSEE at various concentrations (10 - 250 µg/mL) on the activities of ߙ-amylase, ߙ-glucosidase and glucose uptake by yeast cells were used to evaluate its antidiabetic potential. Nitric oxide (NO), 2,2-diphenyl-1-picrylhydrazyl (DPPH) and hydrogen peroxide inhibitory assay were employed to determine its antioxidant effects, while the anti-inflammatory effects were evaluated using human red blood cell (HRBC) membrane stabilization assay. Results: UHSEE inhibited ߙ-amylase and ߙ-glucosidase enzymes but increased glucose uptake by yeast cells in a concentration-dependent manner (p < 0.05). It also inhibited NO, DPPH, hydrogen peroxide and HRBC hemolysis in a concentration-dependent manner (p < 0.05). Conclusion: Uromastyx hardwickii skin exhibits promising good antidiabetic, antioxidant and antiinflammatory properties in vitro. However, its true potentials in this regard needs to be evaluted in vivo.

scholarly journals Molecular Regulation of H3K4 Trimethylation by Wdr82, a Component of Human Set1/COMPASS

2008 ◽  
Vol 28 (24) ◽  
pp. 7337-7344 ◽  
Author(s):  
Min Wu ◽  
Peng Fei Wang ◽  
Jung Shin Lee ◽  
Skylar Martin-Brown ◽  
Laurence Florens ◽  
...  
Keyword(s):  
Posttranslational Modification ◽  
Mammalian Cells ◽  
Molecular Regulation ◽  
Macromolecular Complex ◽  
Dependent Manner ◽  
Specific Component ◽  
Histone H2b ◽  
H3k4 Trimethylation

ABSTRACT In yeast, the macromolecular complex Set1/COMPASS is capable of methylating H3K4, a posttranslational modification associated with actively transcribed genes. There is only one Set1 in yeast; yet in mammalian cells there are multiple H3K4 methylases, including Set1A/B, forming human COMPASS complexes, and MLL1-4, forming human COMPASS-like complexes. We have shown that Wdr82, which associates with chromatin in a histone H2B ubiquitination-dependent manner, is a specific component of Set1 complexes but not that of MLL1-4 complexes. RNA interference-mediated knockdown of Wdr82 results in a reduction in the H3K4 trimethylation levels, although these cells still possess active MLL complexes. Comprehensive in vitro enzymatic studies with Set1 and MLL complexes demonstrated that the Set1 complex is a more robust H3K4 trimethylase in vitro than the MLL complexes. Given our in vivo and in vitro observations, it appears that the human Set1 complex plays a more widespread role in H3K4 trimethylation than do the MLL complexes in mammalian cells.

scholarly journals An intercellular pathway for glucose transport into mouse oocytes

2012 ◽  
Vol 302 (12) ◽  
pp. E1511-E1518 ◽  
Author(s):  
Qiang Wang ◽  
Maggie M. Chi ◽  
Tim Schedl ◽  
Kelle H. Moley
Keyword(s):  
Gap Junctions ◽  
Glucose Uptake ◽  
Glucose Transport ◽  
High Glucose ◽  
Mammalian Cells ◽  
Cumulus Cells ◽  
Oocyte Quality ◽  
Intercellular Pathway

Glucose is an essential nutrient for mammalian cells. Emerging evidence suggests that glucose within the oocyte regulates meiotic maturation. However, it remains controversial as to whether, and if so how, glucose enters oocytes within cumulus-oocyte complexes (COCs). We used a fluorescent glucose derivative (6-NBDG) to trace glucose transport within live mouse COCs and employed inhibitors of glucose transporters (GLUTs) and gap junction proteins to examine their distinct roles in glucose uptake by cumulus cells and the oocyte. We showed that fluorescent glucose enters both cumulus-enclosed and denuded oocytes. Treating COCs with GLUT inhibitors leads to simultaneous decreases in glucose uptake in cumulus cells and the surrounded oocyte but no effect on denuded oocytes. Pharmacological blockade of of gap junctions between the oocyte and cumulus cells significantly inhibited fluorescent glucose transport to oocytes. Moreover, we find that both in vivo hyperglycemic environment and in vitro high-glucose culture increase free glucose levels in oocytes via gap junctional channels. These findings reveal an intercellular pathway for glucose transport into oocytes: glucose is taken up by cumulus cells via the GLUT system and then transferred into the oocyte through gap junctions. This intercellular pathway may partly mediate the effects of high-glucose condition on oocyte quality.

scholarly journals The peroxisome counteracts oxidative stresses by suppressing catalase import via Pex14 phosphorylation

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Kanji Okumoto ◽  
Mahmoud El Shermely ◽  
Masanao Natsui ◽  
Hidetaka Kosako ◽  
Ryuichi Natsuyama ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Protein Import ◽  
Protein Translocation ◽  
Central Component ◽  
Dependent Manner ◽  
Peroxisomal Membrane ◽  
Oxidative Stresses ◽  
Signal Type

Most of peroxisomal matrix proteins including a hydrogen peroxide (H2O2)-decomposing enzyme, catalase, are imported in a peroxisome-targeting signal type-1 (PTS1)-dependent manner. However, little is known about regulation of the membrane-bound protein import machinery. Here, we report that Pex14, a central component of the protein translocation complex in peroxisomal membrane, is phosphorylated in response to oxidative stresses such as H2O2 in mammalian cells. The H2O2-induced phosphorylation of Pex14 at Ser232 suppresses peroxisomal import of catalase in vivo and selectively impairs in vitro the interaction of catalase with the Pex14-Pex5 complex. A phosphomimetic mutant Pex14-S232D elevates the level of cytosolic catalase, but not canonical PTS1-proteins, conferring higher cell resistance to H2O2. We thus suggest that the H2O2-induced phosphorylation of Pex14 spatiotemporally regulates peroxisomal import of catalase, functioning in counteracting action against oxidative stress by the increase of cytosolic catalase.

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