scholarly journals A conserved residue, PomB-F22, in the transmembrane segment of the flagellar stator complex, has a critical role in conducting ions and generating torque

Microbiology ◽  
2011 ◽  
Vol 157 (8) ◽  
pp. 2422-2432 ◽  
Author(s):  
Takashi Terauchi ◽  
Hiroyuki Terashima ◽  
Seiji Kojima ◽  
Michio Homma
Keyword(s):  
Critical Role ◽  
Specific Inhibitor ◽  
Potential Gradient ◽  
Ion Binding ◽  
Conserved Residues ◽  
Flagellar Motors ◽  
Conduction Pathway ◽  
Aspartate Residue ◽  
Conserved Residue ◽  
Hydrophilic Residue

Bacterial flagellar motors exploit the electrochemical potential gradient of a coupling ion (H+ or Na+) as their energy source, and are composed of stator and rotor proteins. Sodium-driven and proton-driven motors have the stator proteins PomA and PomB or MotA and MotB, respectively, which interact with each other in their transmembrane (TM) regions to form an ion channel. The single TM region of PomB or MotB, which forms the ion-conduction pathway together with TM3 and TM4 of PomA or MotA, respectively, has a highly conserved aspartate residue that is the ion binding site and is essential for rotation. To investigate the ion conductivity and selectivity of the Na+-driven PomA/PomB stator complex, we replaced conserved residues predicted to be near the conserved aspartate with H+-type residues, PomA-N194Y, PomB-F22Y and/or PomB-S27T. Motility analysis revealed that the ion specificity was not changed by either of the PomB mutations. PomB-F22Y required a higher concentration of Na+ to exhibit swimming, but this effect was suppressed by additional mutations, PomA-N194Y or PomB-S27T. Moreover, the motility of the PomB-F22Y mutant was resistant to phenamil, a specific inhibitor for the Na+ channel. When PomB-F22 was changed to other amino acids and the effects on swimming ability were investigated, replacement with a hydrophilic residue decreased the maximum swimming speed and conferred strong resistance to phenamil. From these results, we speculate that the Na+ flux is reduced by the PomB-F22Y mutation, and that PomB-F22 is important for the effective release of Na+ from PomB-D24.

Nitric oxide plays a critical role in suppression of T-cell proliferation by mesenchymal stem cells

Blood ◽  
2006 ◽  
Vol 109 (1) ◽  
pp. 228-234 ◽  
Author(s):  
Kazuya Sato ◽  
Katsutoshi Ozaki ◽  
Iekuni Oh ◽  
Akiko Meguro ◽  
Keiko Hatanaka ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Stem Cells ◽  
Cell Proliferation ◽  
T Cells ◽  
Mesenchymal Stem Cells ◽  
T Cell ◽  
Molecular Mechanisms ◽  
Critical Role ◽  
Specific Inhibitor ◽  
T Cell Proliferation

Abstract The molecular mechanisms by which mesenchymal stem cells (MSCs) suppress T-cell proliferation are poorly understood, and whether a soluble factor plays a major role remains controversial. Here we demonstrate that the T-cell–receptor complex is not a target for the suppression, suggesting that downstream signals mediate the suppression. We found that Stat5 phosphorylation in T cells is suppressed in the presence of MSCs and that nitric oxide (NO) is involved in the suppression of Stat5 phosphorylation and T-cell proliferation. The induction of inducible NO synthase (NOS) was readily detected in MSCs but not T cells, and a specific inhibitor of NOS reversed the suppression of Stat5 phosphorylation and T-cell proliferation. This production of NO in the presence of MSCs was mediated by CD4 or CD8 T cells but not by CD19 B cells. Furthermore, inhibitors of prostaglandin synthase or NOS restored the proliferation of T cells, whereas an inhibitor of indoleamine 2,3-dioxygenase and a transforming growth factor–β–neutralizing antibody had no effect. Finally, MSCs from inducible NOS−/− mice had a reduced ability to suppress T-cell proliferation. Taken together, these results suggest that NO produced by MSCs is one of the major mediators of T-cell suppression by MSCs.

Comparison of dynamical features between the fast H-L and the H-I-L transition for EAST RF-heated plasmas

2021 ◽  
Author(s):  
Liang Chen ◽  
Guo Sheng Xu ◽  
Linming Shao ◽  
Wei Gao ◽  
Yifeng Wang ◽  
...  
Keyword(s):  
Temperature Gradient ◽  
Electron Temperature ◽  
Intermediate Phase ◽  
Critical Role ◽  
Potential Gradient ◽  
Flow Shear ◽  
Important Distinction ◽  
Radial Gradient ◽  
Electron Temperature Gradient ◽  
Dynamical Features

Abstract In this paper, a comparison of dynamical features between the fast H-L and the H-I-L transition, which can be identified by the intermediate phase, or ‘I-phase’, has been made for radio-frequency (RF) heated deuterium plasmas in EAST. The fast H-L transition is characterized by a rapid release of stored energy during the transition transient, while the H-I-L transition exhibits a ‘soft’ H-mode termination. One important distinction between the transitions has been observed by dedicated probe measurements slightly inside the separatrix, with respect to the radial gradient of the floating potential, which corresponds to the E×B flow and/or the electron temperature gradient. The potential gradient inside the separatrix oscillates and persists during the stationary I-phase, and shows a larger amplitude than that before the fast H-L transition. The reduction of the gradient leads to the final transition to the L-mode for both the fast H-L and the H-I-L transition. These findings indicate that the mean E×B flow shear and/or edge electron temperature gradient play a critical role underlying the H-L transition physics. In addition, the back transition in EAST is found to be sensitive to magnetic configuration, where the vertical configuration, i.e., inner strike-point located at vertical target, favours access to the H-I-L transition, while the horizontal shape facilitates achievement of the fast H-L transition. The divertor recycling level normalized to electron density is higher before the fast H-L transition, as compared to that before the I-phase, which strongly suggest that the density of the recycled neutrals is an important ingredient in determining the back transition behaviour.

scholarly journals A molecular mechanism of mouse placental spongiotrophoblast differentiation regulated by prolyl oligopeptidase

Zygote ◽  
2019 ◽  
Vol 27 (1) ◽  
pp. 49-53
Author(s):  
Yuki Maruyama ◽  
Atsushi P. Kimura
Keyword(s):  
Molecular Mechanism ◽  
Critical Role ◽  
Specific Inhibitor ◽  
Cell Types ◽  
Giant Cells ◽  
Akt Pathway ◽  
Prolyl Oligopeptidase ◽  
Trophoblast Stem ◽  
Trophoblast Stem Cell ◽  
Trophoblast Giant Cells

SummaryIn eutherian mammals, the placenta plays a critical role in embryo development by supplying nutrients and hormones and mediating interaction with the mother. To establish the fine connection between mother and embryo, the placenta needs to be formed normally, but the mechanism of placental differentiation is not fully understood. We previously revealed that mouse prolyl oligopeptidase (POP) plays a role in trophoblast stem cell (TSC) differentiation into two placental cell types, spongiotrophoblasts (SpT) and trophoblast giant cells. Here, we focused on SpT differentiation and attempted to elucidate a molecular mechanism. ForAscl2,Arnt, andEgfrgenes that are indispensable for SpT formation, we found that a POP-specific inhibitor, SUAM-14746, significantly decreasedAscl2expression, which was consistent with a significant decrease in expression ofFlt1, a gene downstream ofAscl2. Although this downregulation was unlikely to be mediated by the PI3K-Akt pathway, our results indicated that POP controls TSC differentiation into SpT by regulating theAscl2gene.

scholarly journals Phosphorylated AKT preserves stallion sperm viability and motility by inhibiting caspases 3 and 7

Reproduction ◽  
2014 ◽  
Vol 148 (2) ◽  
pp. 221-235 ◽  
Author(s):  
Juan M Gallardo Bolaños ◽  
Carolina M Balao da Silva ◽  
Patricia Martín Muñoz ◽  
Antolín Morillo Rodríguez ◽  
María Plaza Dávila ◽  
...  
Keyword(s):  
Critical Role ◽  
Membrane Integrity ◽  
Specific Inhibitor ◽  
Akt Inhibitor ◽  
Akt Phosphorylation ◽  
Mitochondrial Superoxide ◽  
Phosphorylated Akt ◽  
Sperm Survival

AKT, also referred to as protein kinase B (PKB or RAC), plays a critical role in controlling cell survival and apoptosis. To gain insights into the mechanisms regulating sperm survival after ejaculation, the role of AKT was investigated in stallion spermatozoa using a specific inhibitor and a phosphoflow approach. Stallion spermatozoa were washed and incubated in Biggers–Whitten–Whittingham medium, supplemented with 1% polyvinyl alcohol (PVA) in the presence of 0 (vehicle), 10, 20 or 30 μM SH5, an AKT inhibitor. SH5 treatment reduced the percentage of sperm displaying AKT phosphorylation, with inhibition reaching a maximum after 1 h of incubation. This decrease in phosphorylation was attributable to either dephosphorylation or suppression of the active phosphorylation pathway. Stallion spermatozoa spontaneously dephosphorylated during in vitro incubation, resulting in a lack of a difference in AKT phosphorylation between the SH5-treated sperm and the control after 4 h of incubation. AKT inhibition decreased the proportion of motile spermatozoa (total and progressive) and the sperm velocity. Similarly, AKT inhibition reduced membrane integrity, leading to increased membrane permeability and reduced the mitochondrial membrane potential concomitantly with activation of caspases 3 and 7. However, the percentage of spermatozoa exhibiting oxidative stress, the production of mitochondrial superoxide radicals, DNA oxidation and DNA fragmentation were not affected by AKT inhibition. It is concluded that AKT maintains the membrane integrity of ejaculated stallion spermatozoa, presumably by inhibiting caspases 3 and 7, which prevents the progression of spermatozoa to an incomplete form of apoptosis.Free Spanish abstractA Spanish translation of this abstract is freely available at http://www.reproduction-online.org/content/148/2/221/suppl/DC1.

A critical role for conserved residues in the cleft of HLA-A2 in presentation of a nonapeptide to T cells

Science ◽  
1992 ◽  
Vol 257 (5072) ◽  
pp. 964-967 ◽  
Author(s):  
F Latron ◽  
L Pazmany ◽  
J Morrison ◽  
R Moots ◽  
M. Saper ◽  
...  
Keyword(s):  
T Cells ◽  
Critical Role ◽  
Conserved Residues

Activation of p38 MAP Kinase and JNK But Not ERK Is Required for Erythropoietin-Induced Erythroid Differentiation

Blood ◽  
1998 ◽  
Vol 92 (6) ◽  
pp. 1859-1869 ◽  
Author(s):  
Yuka Nagata ◽  
Noriko Takahashi ◽  
Roger J. Davis ◽  
Kazuo Todokoro
Keyword(s):  
Cell Growth ◽  
Map Kinase ◽  
Antisense Oligonucleotides ◽  
Map Kinases ◽  
Physiological Function ◽  
Critical Role ◽  
Specific Inhibitor ◽  
Erythroid Differentiation ◽  
P38 Map Kinase ◽  
American Society

p38 MAP kinase (p38) and JNK have been described as playing a critical role in the response to a variety of environmental stresses and proinflammatory cytokines. It was recently reported that hematopoietic cytokines activate not only classical MAP kinases (ERK), but also p38 and JNK. However, the physiological function of these kinases in hematopoiesis remains obscure. We found that all MAP kinases examined, ERK1, ERK2, p38, JNK1, and JNK2, were rapidly and transiently activated by erythropoietin (Epo) stimulation in SKT6 cells, which can be induced to differentiate into hemoglobinized cells in response to Epo. Furthermore, p38-specific inhibitor SB203580 but not MEK-specific inhibitor PD98059 significantly suppressed Epo-induced differentiation and antisense oligonucleotides of p38, JNK1, and JNK2, but neither ERK1 nor ERK2 clearly inhibited Epo-induced hemoglobinization. However, in Epo-dependent FD-EPO cells, inhibition of either ERKs, p38, or JNKs suppressed cell growth. Furthermore, forced expression of a gain-of-function MKK6 mutant, which specifically activated p38, induced hemoglobinization of SKT6 cells without Epo. These results indicate that activation of p38 and JNKs but not of ERKs is required for Epo-induced erythroid differentiation of SKT6 cells, whereas all of these kinases are involved in Epo-induced mitogenesis of FD-EPO cells. © 1998 by The American Society of Hematology.

scholarly journals Salt-sensitive hypertension and cardiac hypertrophy in mice deficient in the ubiquitin ligase Nedd4-2

2008 ◽  
Vol 295 (2) ◽  
pp. F462-F470 ◽  
Author(s):  
Peijun P. Shi ◽  
Xiao R. Cao ◽  
Eileen M. Sweezer ◽  
Thomas S. Kinney ◽  
Nathan R. Williams ◽  
...  
Keyword(s):  
Cardiac Hypertrophy ◽  
Critical Role ◽  
Specific Inhibitor ◽  
Normal Diet ◽  
High Salt ◽  
High Salt Diet ◽  
Salt Diet ◽  
Ww Domains ◽  
Salt Sensitive Hypertension

Nedd4-2 has been proposed to play a critical role in regulating epithelial Na+ channel (ENaC) activity. Biochemical and overexpression experiments suggest that Nedd4-2 binds to the PY motifs of ENaC subunits via its WW domains, ubiquitinates them, and decreases their expression on the apical membrane. Phosphorylation of Nedd4-2 (for example by Sgk1) may regulate its binding to ENaC, and thus ENaC ubiquitination. These results suggest that the interaction between Nedd4-2 and ENaC may play a crucial role in Na+ homeostasis and blood pressure (BP) regulation. To test these predictions in vivo, we generated Nedd4-2 null mice. The knockout mice had higher BP on a normal diet and a further increase in BP when on a high-salt diet. The hypertension was probably mediated by ENaC overactivity because 1) Nedd4-2 null mice had higher expression levels of all three ENaC subunits in kidney, but not of other Na+ transporters; 2) the downregulation of ENaC function in colon was impaired; and 3) NaCl-sensitive hypertension was substantially reduced in the presence of amiloride, a specific inhibitor of ENaC. Nedd4-2 null mice on a chronic high-salt diet showed cardiac hypertrophy and markedly depressed cardiac function. Overall, our results demonstrate that in vivo Nedd4-2 is a critical regulator of ENaC activity and BP. The absence of this gene is sufficient to produce salt-sensitive hypertension. This model provides an opportunity to further investigate mechanisms and consequences of this common disorder.

scholarly journals Pgrmc1/BDNF Signaling Plays a Critical Role in Mediating Glia-Neuron Cross Talk

Endocrinology ◽  
2016 ◽  
Vol 157 (5) ◽  
pp. 2067-2079 ◽  
Author(s):  
Fen Sun ◽  
Trinh Nguyen ◽  
Xin Jin ◽  
Renqi Huang ◽  
Zhenglan Chen ◽  
...  
Keyword(s):  
Cross Talk ◽  
Hippocampal Neurons ◽  
Brain Slice ◽  
Critical Role ◽  
Specific Inhibitor ◽  
Neurotrophin Receptor ◽  
Bdnf Signaling ◽  
Brain Slice Cultures ◽  
Enriched Cultures ◽  
Retinoid Acid

Abstract Progesterone (P4) exerts robust cytoprotection in brain slice cultures (containing both neurons and glia), yet such protection is not as evident in neuron-enriched cultures, suggesting that glia may play an indispensable role in P4's neuroprotection. We previously reported that a membrane-associated P4 receptor, P4 receptor membrane component 1, mediates P4-induced brain-derived neurotrophic factor (BDNF) release from glia. Here, we sought to determine whether glia are required for P4's neuroprotection and whether glia's roles are mediated, at least partially, via releasing soluble factors to act on neighboring neurons. Our data demonstrate that P4 increased the level of mature BDNF (neuroprotective) while decreasing pro-BDNF (potentially neurotoxic) in the conditioned media (CMs) of cultured C6 astrocytes. We examined the effects of CMs derived from P4-treated astrocytes (P4-CMs) on 2 neuronal models: 1) all-trans retinoid acid-differentiated SH-SY5Y cells and 2) mouse primary hippocampal neurons. P4-CM increased synaptic marker expression and promoted neuronal survival against H2O2. These effects were attenuated by Y1036 (an inhibitor of neurotrophin receptor [tropomysin-related kinase] signaling), as well as tropomysin-related kinase B-IgG (a more specific inhibitor to block BDNF signaling), which pointed to BDNF as the key protective component within P4-CM. These findings suggest that P4 may exert its maximal protection by triggering a glia-neuron cross talk, in which P4 promotes mature BDNF release from glia to enhance synaptogenesis as well as survival of neurons. This recognition of the importance of glia in mediating P4's neuroprotection may also inform the design of effective therapeutic methods for treating diseases wherein neuronal death and/or synaptic deficits are noted.

scholarly journals Critical role for Epac1 in inflammatory pain controlled by GRK2-mediated phosphorylation of Epac1

2016 ◽  
Vol 113 (11) ◽  
pp. 3036-3041 ◽  
Author(s):  
Pooja Singhmar ◽  
XiaoJiao Huo ◽  
Niels Eijkelkamp ◽  
Susana Rojo Berciano ◽  
Faiza Baameur ◽  
...  
Keyword(s):  
Chronic Pain ◽  
Molecular Mechanism ◽  
Inflammatory Pain ◽  
Critical Role ◽  
Specific Inhibitor ◽  
Mechanical Hyperalgesia ◽  
Dependent Manner ◽  
Phosphorylation Event

cAMP signaling plays a key role in regulating pain sensitivity. Here, we uncover a previously unidentified molecular mechanism in which direct phosphorylation of the exchange protein directly activated by cAMP 1 (EPAC1) by G protein kinase 2 (GRK2) suppresses Epac1-to-Rap1 signaling, thereby inhibiting persistent inflammatory pain. Epac1−/− mice are protected against inflammatory hyperalgesia in the complete Freund’s adjuvant (CFA) model. Moreover, the Epac-specific inhibitor ESI-09 inhibits established CFA-induced mechanical hyperalgesia without affecting normal mechanical sensitivity. At the mechanistic level, CFA increased activity of the Epac target Rap1 in dorsal root ganglia of WT, but not of Epac1−/−, mice. Using sensory neuron-specific overexpression of GRK2 or its kinase-dead mutant in vivo, we demonstrate that GRK2 inhibits CFA-induced hyperalgesia in a kinase activity-dependent manner. In vitro, GRK2 inhibits Epac1-to-Rap1 signaling by phosphorylation of Epac1 at Ser-108 in the Disheveled/Egl-10/pleckstrin domain. This phosphorylation event inhibits agonist-induced translocation of Epac1 to the plasma membrane, thereby reducing Rap1 activation. Finally, we show that GRK2 inhibits Epac1-mediated sensitization of the mechanosensor Piezo2 and that Piezo2 contributes to inflammatory mechanical hyperalgesia. Collectively, these findings identify a key role of Epac1 in chronic inflammatory pain and a molecular mechanism for controlling Epac1 activity and chronic pain through phosphorylation of Epac1 at Ser-108. Importantly, using the Epac inhibitor ESI-09, we validate Epac1 as a potential therapeutic target for chronic pain.

Prediction of the Tertiary Structure of α-Glycosidase from Aspergillus niger by Homology Modeling

2011 ◽  
Vol 399-401 ◽  
pp. 2160-2163
Author(s):  
Fa Xiang Wang ◽  
Qin Yun Wang ◽  
Yong Le Liu ◽  
Jian Yu
Keyword(s):  
Homology Modeling ◽  
Tertiary Structure ◽  
Catalytic Domain ◽  
Molecular Model ◽  
Critical Role ◽  
Three Dimensional ◽  
Primary Metabolism ◽  
Structural Domains ◽  
Conserved Residues ◽  
Substrate Binding Sites

α-Glucosidases play critical role both in primary metabolism and in glycoconjugate biosynthesis and processing. In this paper, the reasonable three-dimensional molecular model of AglA was generated by homology modeling. This modeled protein is divided into five major structural domains, and the catalytic domain is classical (β/α) 8 barrel with the active site pocket positioned at its C-terminal side. With analyses of conserved residues and overlay of homology structures, the residues Tyr 662, Tyr527, Glu521, His238 and Tyr235 was predicted as the main substrate binding sites, and residues Asp490, Glu493 and Asp660 were deduced to be the acid/base catalytic residues.

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