scholarly journals Cyclic AMP-dependent Protein Lysine Acylation in Mycobacteria Regulates Fatty Acid and Propionate Metabolism

2013 ◽  
Vol 288 (20) ◽  
pp. 14114-14124 ◽  
Author(s):  
Subhalaxmi Nambi ◽  
Kallol Gupta ◽  
Moitrayee Bhattacharyya ◽  
Parvathy Ramakrishnan ◽  
Vaishnavi Ravikumar ◽  
...  
Keyword(s):  
Critical Role ◽  
Fatty Acyl ◽  
Intracellular Camp ◽  
Dependent Manner ◽  
Multiple Substrates ◽  
Propionate Metabolism ◽  
Lysine Residues ◽  
Dependent Protein ◽  
First Time ◽  
Camp Levels

Acetylation of lysine residues is a posttranslational modification that is used by both eukaryotes and prokaryotes to regulate a variety of biological processes. Here we identify multiple substrates for the cAMP-dependent protein lysine acetyltransferase from Mycobacterium tuberculosis (KATmt). We demonstrate that a catalytically important lysine residue in a number of FadD (fatty acyl CoA synthetase) enzymes is acetylated by KATmt in a cAMP-dependent manner and that acetylation inhibits the activity of FadD enzymes. A sirtuin-like enzyme can deacetylate multiple FadDs, thus completing the regulatory cycle. Using a strain deleted for the KATmt ortholog in Mycobacterium bovis Bacillus Calmette-Guérin (BCG), we show for the first time that acetylation is dependent on intracellular cAMP levels. KATmt can utilize propionyl CoA as a substrate and, therefore, plays a critical role in alleviating propionyl CoA toxicity in mycobacteria by inactivating acyl CoA synthetase (ACS). The precision by which mycobacteria can regulate the metabolism of fatty acids in a cAMP-dependent manner appears to be unparalleled in other biological organisms and is ideally suited to adapt to the complex environment that pathogenic mycobacteria experience in the host.

VIP inhibits basal and histamine-stimulated proliferation of human airway smooth muscle cells

1995 ◽  
Vol 268 (6) ◽  
pp. L1047-L1051 ◽  
Author(s):  
K. Maruno ◽  
A. Absood ◽  
S. I. Said
Keyword(s):  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Thymidine Incorporation ◽  
Inhibitory Effect ◽  
Airway Smooth Muscle Cells ◽  
Dependent Manner ◽  
Cyclic Monophosphate ◽  
Cell Counts ◽  
Dependent Protein ◽  
Camp Levels

Airway smooth muscle (ASM) cell proliferation contributes to increased airway resistance in bronchial asthma. We have examined the modulation of ASM proliferation by vasoactive intestinal peptide (VIP), a cotransmitter of airway relaxation. Human ASM cells were grown in culture as a monolayer. VIP (1.0 nM-1.0 microM) inhibited proliferation in a dose-dependent manner by up to 82% on day 2, but the related peptide glucagon had no effect. Histamine (100 nM-100 microM) increased cell counts by 66%, but in the presence of VIP, cell counts and [3H]thymidine incorporation were reduced by up to 55%. Adenosine 3',5'-cyclic monophosphate (cAMP)-promoting agents, including 3-isobutyl-1-methylxanthine, forskolin, and 8-bromo-adenosine 3',5'-cyclic monophosphate, alone and especially combined with VIP, reduced cell counts and [3H]thymidine incorporation, in correlation with cAMP levels. KT-5720 (1.0 nM-1.0 microM), a selective inhibitor of cAMP-dependent protein kinase A (PKA), abolished the inhibitory effect of VIP. The results show that VIP selectively and potently inhibits human ASM cell growth and multiplication, and nullifies the mitogenic effect of histamine, by a PKA-mediated mechanism. A deficiency of VIP may lead to ASM hyperplasia due to unopposed stimulation by endogenous mitogens.

scholarly journals miR-103 Regulates Oxidative Stress by Targeting the BCL2/Adenovirus E1B 19 kDa Interacting Protein 3 in HUVECs

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Mao-Chun Xu ◽  
Xiu-Fang Gao ◽  
Changwu Ruan ◽  
Zhi-Ru Ge ◽  
Ji-De Lu ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Critical Role ◽  
Cell Activity ◽  
Antioxidant Effect ◽  
Dependent Manner ◽  
Ros Production ◽  
Interacting Protein ◽  
Adenovirus E1b ◽  
Antioxidative Therapy ◽  
First Time

Oxidative stress plays a critical role in cardiovascular diseases. Salidroside, a glycoside fromRhodiola rosea, has been used as an antioxidative therapy for oxidative injury in cardiac diseases. However, the mechanism underlying its antioxidant effect needs to be elucidated. Treatment of HUVECs with H2O2significantly decreased the expression of miR-103 in a dose- and time-dependent manner, whereas pretreatment with salidroside significantly inhibited this decrease. Subsequent analysis showed that overexpression of miR-103 abrogated cell activity and ROS production induced by H2O2. Bcl2/adenovirus E1B 19 kDa interacting protein 3 (BNIP3) was determined to be a novel miR-103 target in HUVECs. Interestingly, H2O2treatment upregulated BNIP3 expression; in turn, this effect was inhibited by pretreatment with salidroside. Further studies confirmed that the knockdown of BNIP3 enhanced cell activity and suppressed the ROS production induced by H2O2. These results demonstrated for the first time that salidroside protects HUVECs in part by upregulating the expression of miR-103, which mediates BNIP3 downregulation and plays an important role in the cytoprotective actions.

scholarly journals Borrelia burgdorferi Infection-Associated Surface Proteins ErpP, ErpA, and ErpC Bind Human Plasminogen

2008 ◽  
Vol 77 (1) ◽  
pp. 300-306 ◽  
Author(s):  
Catherine A. Brissette ◽  
Katrin Haupt ◽  
Diana Barthel ◽  
Anne E. Cooley ◽  
Amy Bowman ◽  
...  
Keyword(s):  
Borrelia Burgdorferi ◽  
Critical Role ◽  
Aminocaproic Acid ◽  
Factor H ◽  
Surface Proteins ◽  
Primary Role ◽  
Dependent Manner ◽  
Lysine Residues ◽  
Complement Regulator ◽  
Mammalian Infection

ABSTRACT Host-derived plasmin plays a critical role in mammalian infection by Borrelia burgdorferi. The Lyme disease spirochete expresses several plasminogen-binding proteins. Bound plasminogen is converted to the serine protease plasmin and thereby may facilitate the bacterium's dissemination throughout the host by degrading extracellular matrix. In this work, we demonstrate plasminogen binding by three highly similar borrelial outer surface proteins, ErpP, ErpA, and ErpC, all of which are expressed during mammalian infection. Extensive characterization of ErpP demonstrated that this protein bound in a dose-dependent manner to lysine binding site I of plasminogen. Removal of three lysine residues from the carboxy terminus of ErpP significantly reduced binding of plasminogen, and the presence of a lysine analog, ε-aminocaproic acid, inhibited the ErpP-plasminogen interaction, thus strongly pointing to a primary role for lysine residues in plasminogen binding. Ionic interactions are not required in ErpP binding of plasminogen, as addition of excess NaCl or the polyanion heparin did not have any significant effect on binding. Plasminogen bound to ErpP could be converted to the active enzyme, plasmin. The three plasminogen-binding Erp proteins can also bind the host complement regulator factor H. Plasminogen and factor H bound simultaneously and did not compete for binding to ErpP, indicating separate binding sites for both host ligands and the ability of the borrelial surface proteins to bind both host proteins.

scholarly journals Gi proteins mediate activation of the canonical hedgehog pathway in the myocardium

2014 ◽  
Vol 307 (1) ◽  
pp. H66-H72 ◽  
Author(s):  
Christian J. Carbe ◽  
Lan Cheng ◽  
Sankar Addya ◽  
Jessica I. Gold ◽  
Erhe Gao ◽  
...  
Keyword(s):  
Purinergic Signaling ◽  
Transmembrane Protein ◽  
Critical Role ◽  
Transcriptional Response ◽  
Neonatal Rat ◽  
Small Subset ◽  
Dependent Manner ◽  
Gli1 Expression ◽  
Ventricular Cardiomyocytes ◽  
First Time

During myocardial ischemia, upregulation of the hedgehog (Hh) pathway promotes neovascularization and increases cardiomyocyte survival. The canonical Hh pathway activates a transcriptional program through the Gli family of transcription factors by derepression of the seven-transmembrane protein smoothened (Smo). The mechanisms linking Smo to Gli are complex and, in some cell types, involve coupling of Smo to Gi proteins. In the present study, we investigated, for the first time, the transcriptional response of cardiomyocytes to sonic hedgehog (Shh) and the role of Gi protein utilization. Our results show that Shh strongly activates Gli1 expression by quantitative PCR in a Smo-dependent manner in neonatal rat ventricular cardiomyocytes. Microarray analysis of gene expression changes elicited by Shh and sensitive to a Smo inhibitor identified a small subset of 37 cardiomyocyte-specific genes regulated by Shh, including some in the PKA and purinergic signaling pathways. In addition, neonatal rat ventricular cardiomyocytes infected with an adenovirus encoding GiCT, a peptide that impairs receptor-Gi protein coupling, showed reduced activation of Hh targets. In vitro data were confirmed in transgenic mice with cardiomyocyte-inducible GiCT expression. Transgenic GiCT mice showed specific reduction of Gli1 expression in the heart under basal conditions and failed to upregulate the Hh pathway upon ischemia and reperfusion injury, unlike their littermate controls. This study characterizes, for the first time, the transcriptional response of cardiomyocytes to Shh and establishes a critical role for Smo coupling to Gi in Hh signaling in the normal and ischemic myocardium.

Phosphorylation of myristoylated alanine-rich C kinase substrate is involved in the cAMP-dependent amylase release in parotid acinar cells

2009 ◽  
Vol 296 (6) ◽  
pp. G1382-G1390 ◽  
Author(s):  
Keitaro Satoh ◽  
Miwako Matsuki-Fukushima ◽  
Bing Qi ◽  
Ming-Yu Guo ◽  
Takanori Narita ◽  
...  
Keyword(s):  
Protein Kinase ◽  
Acinar Cells ◽  
Cellular Migration ◽  
Intracellular Camp ◽  
Dependent Manner ◽  
Amylase Release ◽  
Kinase Substrate ◽  
Parotid Acinar Cells ◽  
Dependent Protein ◽  
Rat Parotid

Myristoylated alanine-rich C kinase substrate (MARCKS) is known as a major cellular substrate for protein kinase C (PKC). MARCKS has been implicated in the regulation of brain development and postnatal survival, cellular migration and adhesion, as well as phagocytosis, endocytosis, and exocytosis. The involvement of MARCKS phosphorylation in secretory function has been reported in Ca2+-mediated exocytosis. In rat parotid acinar cells, the activation of β-adrenergic receptors provokes exocytotic amylase release via accumulation of intracellular cAMP levels. Here, we studied the involvement of MARCKS phosphorylation in the cAMP-dependent amylase release in rat parotid acinar cells. MARCKS protein was detected in rat parotid acinar cells by Western blotting. The β-adrenergic agonist isoproterenol (IPR) induced MARCKS phosphorylation in a time-dependent manner. Translocation of a part of phosphorylated MARCKS from the membrane to the cytosol and enhancement of MARCKS phosphorylation at the apical membrane site induced by IPR were observed by immunohistochemistry. H89, a cAMP-dependent protein kinase (PKA) inhibitor, inhibited the IPR-induced MARCKS phosphorylation. The PKCδ inhibitor rottlerin inhibited the IPR-induced MARCKS phosphorylation and amylase release. IPR activated PKCδ, and the effects of IPR were inhibited by the PKA inhibitors. A MARCKS-related peptide partially inhibited the IPR-induced amylase release. These findings suggest that MARCKS phosphorylation via the activation of PKCδ, which is downstream of PKA activation, is involved in the cAMP-dependent amylase release in parotid acinar cells.

scholarly journals WNT/β-CATENIN modulates the axial identity of ES derived human neural crest

2019 ◽  
Author(s):  
Gustavo A. Gomez ◽  
Maneeshi S. Prasad ◽  
Man Wong ◽  
Rebekah M. Charney ◽  
Patrick B. Shelar ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Neural Crest ◽  
Critical Role ◽  
Embryonic Stem ◽  
Dependent Manner ◽  
Robust Model ◽  
Magnitude Variation ◽  
First Time ◽  
Dose Dependent ◽  
Wnt Signal

ABSTRACTThe WNT/β-CATENIN pathway is critical for neural crest (NC) formation. However, the effects of the magnitude of the signal remains poorly defined. Here we evaluate the consequences of WNT magnitude variation in a robust model of human NC formation. This model is based on human embryonic stem cells induced by WNT signaling through the small molecule CHIR9902. In addition to its known effect on NC formation, we find that the WNT signal modulates the anterior-posterior axial identity of NCCs in a dose dependent manner, with low WNT leading to anterior OTX+, HOX-NC, and high WNT leading to posterior OTX−, HOX+ NC. Differentiation tests of posterior NC confirm expected derivatives including posterior specific adrenal derivatives, and display partial capacity to generate anterior ectomesenchymal derivatives. Furthermore, unlike anterior NC, posterior NC transit through a TBXT+/SOX2+ neuromesodermal precursor-like intermediate. Finally, we analyze the contributions of other signaling pathways in posterior NC formation, and suggest a critical role for FGF in survival/proliferation, and a requirement of BMP for NC maturation. As expected RA and FGF are able to modulate HOX expression in the posterior NC, but surprisingly, RA supplementation prohibits anterior, but only reduces, posterior NC formation. This work reveals for the first time that the amplitude of WNT signaling can modulate the axial identity of NC cells in humans.

scholarly journals Chlorin e6 Prevents ADP-Induced Platelet Aggregation by Decreasing PI3K-Akt Phosphorylation and Promoting cAMP Production

2013 ◽  
Vol 2013 ◽  
pp. 1-11 ◽  
Author(s):  
Ji Young Park ◽  
Hyun Dong Ji ◽  
Bo Ra Jeon ◽  
Eun Ju Im ◽  
Young Min Son ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Underlying Mechanism ◽  
Serotonin Release ◽  
Intracellular Camp ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Akt Phosphorylation ◽  
Mitogen Activated Protein ◽  
Induced Aggregation ◽  
Camp Levels

A number of reagents that prevent thrombosis have been developed but were found to have serious side effects. Therefore, we sought to identify complementary and alternative medicinal materials that are safe and have long-term efficacy. In the present studies, we have assessed the ability of chlorine e6 (CE6) to inhibit ADP-induced aggregation of rat platelets and elucidated the underlying mechanism. CE6 inhibited platelet aggregation induced by 10 µM ADP in a concentration-dependent manner and decreased intracellular calcium mobilization and granule secretion (i.e., ATP and serotonin release). Western blotting revealed that CE6 strongly inhibited the phosphorylations of PI3K, Akt, c-Jun N-terminal kinase (JNK), and different mitogen-activated protein kinases (MAPKs) including extracellular signal-regulated kinase 1/2 (ERK1/2) as well as p38-MAPK. Our study also demonstrated that CE6 significantly elevated intracellular cAMP levels and decreased thromboxane A2formation in a concentration-dependent manner. Furthermore, we determined that CE6 initiated the activation of PKA, an effector of cAMP. Taken together, our findings indicate that CE6 may inhibit ADP-induced platelet activation by elevating cAMP levels and suppressing PI3K/Akt activity. Finally, these results suggest that CE6 could be developed as therapeutic agent that helps prevent thrombosis and ischemia.

REGULATION OF CYCLIC NUCLEOTIDE PHOSPHODIESTERASE ACTIVITY IN PLATELETS BY PHOSPHORYLATION

1987 ◽  
Author(s):  
P G Grant ◽  
A F Mannarino ◽  
R W Colman
Keyword(s):  
Protein Kinase ◽  
Cyclic Nucleotide ◽  
Kinase Inhibitor ◽  
Specific Activity ◽  
Intracellular Camp ◽  
Blue Dextran ◽  
Dependent Protein ◽  
Camp Levels ◽  
Hydrolysis Of ◽  
Stimulation Of

Cyclic nucleotide phosphodiesterases (PDE) provide the only known pathway for the hydrolysis of cyclic nucleotides in cells and thus have the potential for modulating the effects of cAMP and cGMP on cells. In platelets a rise in intracellular cAMP levels inhibits platelet aggregation and secretion. Since cAMP exerts many of its effects through a cAMP-dependent kinase we questioned whether phosphorylation of cAMP PDE might be a mode for regulation of PDE activity in platelets. When platelets were incubated for 10 min with forskolin (100 μM) the level of cAMP rose at least 10-fold.When the low Km cyclic nucleotide PDE was isolated from freeze-thaw lysates of forskolin treated platelets by chromatography on blue dextran-Sepharose, the specific activity of this enzyme was increased 3 to 13-fold over similarly processed control platelets. The specific activity of a second PDE, the cGMP-stimulated cAMP PDE, was increased 1.5 to 3-fold by forskolin treatment of platelets. Forskolin had no direct effect on either purified PDE. The stimulation of the low Km cAMP PDE activity by exposure of platelets to forskolin was blocked when the platelets were simultaneously treated with the protein kinase inhibitor H-8 (100 μM) which is most potent toward cAMP dependent protein kinase indicating that this kinase may be responsible for the stimulation. When platelets which had been prelabeled with 32P inorganic phosphate were treated with forskolin and the low Km cAMP PDE isolated by blue dextran-Sepharose chromatography, a protein migrating in SDS gels at Mr=110,000, the molecular weight of the low Km cAMP PDE, was labeled indicating that phosphorylation of the PDE occurred coincident with stimulation of activity. These results suggest that phosphorylation of the low Km cAMP PDE by protein kinase may be an important regulatory mechanism for cAMP PDE activity and cyclic nucleotide levels in platelets.

The effects of GH-releasing peptide-6 (GHRP-6) and GHRP-2 on intracellular adenosine 3′,5′-monophosphate (cAMP) levels and GH secretion in ovine and rat somatotrophs

1996 ◽  
Vol 148 (2) ◽  
pp. 197-205 ◽  
Author(s):  
D Wu ◽  
C Chen ◽  
J Zhang ◽  
C Y Bowers ◽  
I J Clarke
Keyword(s):  
Additive Effect ◽  
Intracellular Camp ◽  
Pituitary Cells ◽  
Dependent Manner ◽  
Camp Accumulation ◽  
Gh Secretion ◽  
Rat Pituitary ◽  
Rat Pituitary Cells ◽  
Camp Levels

Abstract The mechanism of action of GH-releasing peptide-6 (GHRP-6) and GHRP-2 on GH release was investigated in ovine and rat pituitary cells in vitro. In partially purified sheep somatotrophs, GHRP-2 and GH-releasing factor (GRF) increased intracellular cyclic AMP (cAMP) concentrations and caused GH release in a dose-dependent manner; GHRP-6 did not increase cAMP levels. An additive effect of maximal doses of GRF and GHRP-2 was observed in both cAMP and GH levels whereas combined GHRP-6 and GHRP-2 at maximal doses produced an additive effect on GH release only. Pretreatment of the cells with MDL 12,330A, an adenylyl cyclase inhibitor, prevented cAMP accumulation and the subsequent release of GH that was caused by either GHRP-2 or GRF. The cAMP antagonist, Rp-cAMP also blocked GH release in response to GHRP-2 and GRF. The cAMP antagonist did not prevent the effect of GHRP-6 on GH secretion whereas MDL 12,330A partially reduced the effect. An antagonist for the GRF receptor, [Ac-Tyr1,d-Arg2]-GRF 1–29, significantly diminished the effect of GHRP-2 and GRF on cAMP accumulation and GH release, but did not affect GH release induced by GHRP-6. Somatostatin prevented cAMP accumulation and GH release responses to GHRP-2, GRF and GHRP-6. Ca2+ channel blockade did not affect the cAMP increase in response to GHRP-2 or GRF but totally prevented GH release in response to GHRP-2, GRF and GHRP-6. These results indicated that GHRP-2 acts on ovine pituitary somatotrophs to increase cAMP concentration in a manner similar to that of GRF; this occurs even during the blockade of Ca2+ influx. GHRP-6 caused GH release without an increase in intracellular cAMP levels. GH release in response to all three secretagogues was reduced by somatostatin and was dependent upon the influx of extracellular Ca2+. The additive effect of GHRP-2 and GRF or GHRP-6 suggested that the three peptides may act on different receptors. In rat pituitary cell cultures, GHRP-6 had no effect on cAMP levels, but potentiated the effect of GRF on cAMP accumulation. The synergistic effect of GRF and GHRP-6 on cAMP accumulation did not occur in sheep somatotrophs. Whereas GHRP-2 caused cAMP accumulation in sheep somatotrophs, it did not do so in rat pituitary cells. These data indicate species differences in the response of pituitary somatotrophs to the GHRPs and this is probably due to different subtypes of GHRP receptor in rat or sheep. Journal of Endocrinology (1996) 148, 197–205

scholarly journals Elevated intracellular cAMP concentration mediates growth suppression in glioma cells

2019 ◽  
Author(s):  
Dewi Safitri ◽  
Harriet Potter ◽  
Matthew Harris ◽  
Ian Winfield ◽  
Liliya Kopanitsa ◽  
...  
Keyword(s):  
Cell Growth ◽  
Phase Cell ◽  
Intracellular Camp ◽  
Dependent Manner ◽  
Camp Concentration ◽  
Cycle Arrest ◽  
Pde Inhibitors ◽  
M Phase ◽  
Camp Levels ◽  
Intracellular Camp Concentration

SUMMARYSupressed levels of intracellular cAMP have been associated with malignancy. Thus, elevating cAMP through activation of adenylyl cyclase (AC) or by inhibition of phosphodiesterase (PDE) may be therapeutically beneficial. Here, we demonstrate that elevated cAMP levels suppress growth in C6 cells (a model of glioma) through treatment with forskolin, an AC activator, or a range of small molecule PDE inhibitors with differing selectivity profiles. Forskolin suppressed cell growth in a protein kinase A (PKA)-dependent manner by inducing a G2/M phase cell cycle arrest. In contrast, trequinsin (a non-selective PDE2/3/7 inhibitor), not only inhibited cell growth via PKA, but also stimulated (independent of PKA) caspase-3/-7 and induced an aneuploidy phenotype. Interestingly, a cocktail of individual PDE 2,3,7 inhibitors suppressed cell growth in a manner analogous to forskolin but not trequinsin. Finally, we demonstrate that concomitant targeting of both AC and PDEs synergistically elevated intracellular cAMP levels thereby potentiating their antiproliferative actions.

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