scholarly journals Contribution of Val/Ile87 residue in the extracellular domain in agonist-induced current responses of the human and rat P2X7 receptors

2020 ◽  
Author(s):  
Emily A Caseley ◽  
Stephen P Muench ◽  
Lin-Hua Jiang
Keyword(s):  
P2x7 Receptor ◽  
Mammalian Species ◽  
Critical Role ◽  
Extracellular Domain ◽  
Cation Channel ◽  
Induced Current ◽  
P2x7 Receptors ◽  
Functional Differences ◽  
Dependent Function ◽  
Molecular Determinants

Abstract The P2X7 receptor (P2X7R) is an ATP-gated cation channel with a critical role in many physiological and pathological processes, and shows prominent functional differences across mammalian species, exemplified by larger current responses of the rat (r) P2X7R to ATP and its analogue BzATP and a greater sensitivity to agonists compared with the human (h) P2X7R. Here, we showed that substitution of Val87 residue in the extracellular domain of the hP2X7R with isoleucine in the rP2X7R increased the current responses of the hP2X7R to both ATP and BzATP. Conversely, introduction of reciprocal I87V mutation in the rP2X7R led to a noticeable but statistically insignificant reduction in the current responses of the rP2X7R to ATP and BzATP. The mutations did not affect the sensitivity of the human and rat P2X7Rs to ATP and BzATP. These results suggest a contribution of Val/Ile87 in agonist-induced current responses of human and rat P2X7Rs, which helps to better understand the molecular determinants for species-dependent function of the mammalian P2X7Rs.

scholarly journals Functional Coupling between the P2X7 Receptor and Pannexin-1 Channel in Rat Trigeminal Ganglion Neurons

2021 ◽  
Vol 22 (11) ◽  
pp. 5978
Author(s):  
Hiroyuki Inoue ◽  
Hidetaka Kuroda ◽  
Wataru Ofusa ◽  
Sadao Oyama ◽  
Maki Kimura ◽  
...  
Keyword(s):  
Trigeminal Ganglion ◽  
P2x7 Receptor ◽  
P2x Receptor ◽  
Functional Coupling ◽  
Dependent Manner ◽  
High Concentration ◽  
Receptor Antagonists ◽  
P2x7 Receptors ◽  
Pannexin 1 ◽  
Ganglion Neurons

The ionotropic P2X receptor, P2X7, is believed to regulate and/or generate nociceptive pain, and pain in several neuropathological diseases. Although there is a known relationship between P2X7 receptor activity and pain sensing, its detailed functional properties in trigeminal ganglion (TG) neurons remains unclear. We examined the electrophysiological and pharmacological characteristics of the P2X7 receptor and its functional coupling with other P2X receptors and pannexin-1 (PANX1) channels in primary cultured rat TG neurons, using whole-cell patch-clamp recordings. Application of ATP and Bz-ATP induced long-lasting biphasic inward currents that were more sensitive to extracellular Bz-ATP than ATP, indicating that the current was carried by P2X7 receptors. While the biphasic current densities of the first and second components were increased by Bz-ATP in a concentration dependent manner; current duration was only affected in the second component. These currents were significantly inhibited by P2X7 receptor antagonists, while only the second component was inhibited by P2X1, 3, and 4 receptor antagonists, PANX1 channel inhibitors, and extracellular ATPase. Taken together, our data suggests that autocrine or paracrine signaling via the P2X7-PANX1-P2X receptor/channel complex may play important roles in several pain sensing pathways via long-lasting neuronal activity driven by extracellular high-concentration ATP following tissue damage in the orofacial area.

scholarly journals Kinase CDK2 in Mammalian Meiotic Prophase I: Screening for Hetero- and Homomorphic Sex Chromosomes

2021 ◽  
Vol 22 (4) ◽  
pp. 1969
Author(s):  
Sergey Matveevsky ◽  
Tsenka Chassovnikarova ◽  
Tatiana Grishaeva ◽  
Maret Atsaeva ◽  
Vasilii Malygin ◽  
...  
Keyword(s):  
Sex Chromosomes ◽  
Mammalian Species ◽  
Critical Role ◽  
Eukaryotic Cell ◽  
Repair System ◽  
Prophase I ◽  
Meiotic Chromosomes ◽  
Y Chromosomes ◽  
Dna Mismatch Repair System ◽  
Male Sex

Cyclin-dependent kinases (CDKs) are crucial regulators of the eukaryotic cell cycle. The critical role of CDK2 in the progression of meiosis was demonstrated in a single mammalian species, the mouse. We used immunocytochemistry to study the localization of CDK2 during meiosis in seven rodent species that possess hetero- and homomorphic male sex chromosomes. To compare the distribution of CDK2 in XY and XX male sex chromosomes, we performed multi-round immunostaining of a number of marker proteins in meiotic chromosomes of the rat and subterranean mole voles. Antibodies to the following proteins were used: RAD51, a member of the double-stranded DNA break repair machinery; MLH1, a component of the DNA mismatch repair system; and SUN1, which is involved in the connection between the meiotic telomeres and nuclear envelope, alongside the synaptic protein SYCP3 and kinetochore marker CREST. Using an enhanced protocol, we were able to assess the distribution of as many as four separate proteins in the same meiotic cell. We showed that during prophase I, CDK2 localizes to telomeric and interstitial regions of autosomes in all species investigated (rat, vole, hamster, subterranean mole voles, and mole rats). In sex bivalents following synaptic specificity, the CDK2 signals were distributed in three different modes. In the XY bivalent in the rat and mole rat, we detected numerous CDK2 signals in asynaptic regions and a single CDK2 focus on synaptic segments, similar to the mouse sex chromosomes. In the mole voles, which have unique XX sex chromosomes in males, CDK2 signals were nevertheless distributed similarly to the rat XY sex chromosomes. In the vole, sex chromosomes did not synapse, but demonstrated CDK2 signals of varying intensity, similar to the rat X and Y chromosomes. In female mole voles, the XX bivalent had CDK2 pattern similar to autosomes of all species. In the hamster, CDK2 signals were revealed in telomeric regions in the short synaptic segment of the sex bivalent. We found that CDK2 signals colocalize with SUN1 and MLH1 signals in meiotic chromosomes in rats and mole voles, similar to the mouse. The difference in CDK2 manifestation at the prophase I sex chromosomes can be considered an example of the rapid chromosome evolution in mammals.

scholarly journals Inhibitory Effects of Metals on ATP-Induced Current Through P2X7 Receptor in NG108-15 Cells

2002 ◽  
Vol 89 (3) ◽  
pp. 296-301 ◽  
Author(s):  
Tomokazu Watano ◽  
Isao Matsuoka ◽  
Junko Kimura
Keyword(s):  
P2x7 Receptor ◽  
Induced Current ◽  
Inhibitory Effects

scholarly journals Selective interactions among the multiple connexin proteins expressed in the vertebrate lens: the second extracellular domain is a determinant of compatibility between connexins.

1994 ◽  
Vol 125 (4) ◽  
pp. 879-892 ◽  
Author(s):  
T W White ◽  
R Bruzzone ◽  
S Wolfram ◽  
D L Paul ◽  
D A Goodenough
Keyword(s):  
Extracellular Domain ◽  
Adjacent Cell ◽  
Molecular Determinants ◽  
Voltage Dependent ◽  
Cytoplasmic Acidification ◽  
Intercellular Channels ◽  
Connexin Proteins ◽  
Heterotypic Channels ◽  
Selective Process ◽  
High Conductance

Gap junctions are collections of intercellular channels composed of structural proteins called connexins (Cx). We have examined the functional interactions of the three rodent connexins present in the lens, Cx43, Cx46, and Cx50, by expressing them in paired Xenopus oocytes. Homotypic channels containing Cx43, Cx46, or Cx50 all developed high conductance. heterotypic channels composed of Cx46 paired with either Cx43 or Cx50 were also well coupled, whereas Cx50 did not form functional channels with Cx43. We also examined the functional response of homotypic and heterotypic channels to transjunctional voltage and cytoplasmic acidification. We show that all lens connexins exhibited sensitivity to cytoplasmic acidification as well as to voltage, and that voltage-dependent closure of heterotypic channels for a given connexin was dramatically influenced by its partner connexins in the adjacent cell. Based on the observation that Cx43 can discriminate between Cx46 and Cx50, we investigated the molecular determinants that specify compatibility by constructing chimeric connexins from portions of Cx46 and Cx50 and testing them for their ability to form channels with Cx43. When the second extracellular (E2) domain in Cx46 was replaced with the E2 of Cx50, the resulting chimera could no longer form heterotypic channels with Cx43. A reciprocal chimera, where the E2 of Cx46 was inserted into Cx50, acquired the ability to functionally interact with Cx43. Together, these results demonstrate that formation of intercellular channels is a selective process dependent on the identity of the connexins expressed in adjacent cells, and that the second extracellular domain is a determinant of heterotypic compatibility between connexins.

scholarly journals RNF216 is essential for spermatogenesis and male fertility†

2019 ◽  
Vol 100 (5) ◽  
pp. 1132-1134 ◽  
Author(s):  
Ashley F Melnick ◽  
Yuen Gao ◽  
Jiali Liu ◽  
Deqiang Ding ◽  
Alicia Predom ◽  
...  
Keyword(s):  
Male Fertility ◽  
Mammalian Species ◽  
Critical Role ◽  
Ring Finger ◽  
Gonadal Development ◽  
Cellular Protein ◽  
Ring Finger Protein ◽  
Targeted Deletion ◽  
Finger Protein ◽  
Essential Function

Abstract Ring finger protein 216 (RNF216) belongs to the RING family of E3 ubiquitin ligases that are involved in cellular protein degradation. Mutations in human Rnf216 gene have been identified in Gordon Holmes syndrome, which is defined by ataxia, dementia, and hypogonadotropism. However, the gene function of Rnf216 in mammalian species remains unknown. Here, we show that targeted deletion of Rnf216 in mice results in disruption in spermatogenesis and male infertility. RNF216 is not required for female fertility. These findings reveal an essential function of RNF216 in spermatogenesis and male fertility and suggest a critical role for RNF216 in human gonadal development.

Adenosine Triphosphate–Induced Shedding of CD23 and L-Selectin (CD62L) From Lymphocytes Is Mediated by the Same Receptor but Different Metalloproteases

Blood ◽  
1998 ◽  
Vol 92 (3) ◽  
pp. 946-951 ◽  
Author(s):  
B. Gu ◽  
L.J. Bendall ◽  
J.S. Wiley
Keyword(s):  
Adenosine Triphosphate ◽  
Phorbol Ester ◽  
P2x7 Receptor ◽  
Transmembrane Protein ◽  
Lymphocytic Leukemia ◽  
P2x7 Receptors ◽  
Extracellular Adenosine ◽  
Zinc Chelator ◽  
High Concentrations ◽  
American Society

CD23 is a transmembrane protein expressed on the surface of B-lymphocytes that binds IgE, CD21, CD11b, and CD11c. High concentrations of soluble CD23 and L-selectin are found in the serum of patients with B-chronic lymphocytic leukemia (B-CLL). Because extracellular adenosine triphosphate (ATP) causes shedding of L-selectin via activation of P2Z/P2X7 receptors expressed on B-CLL lymphocytes, we studied the effect of ATP on shedding of CD23. ATP-induced shedding of CD23 at an initial rate of 12% of that for L-selectin, whereas the EC50 for ATP was identical (35 μmol/L) for shedding of both molecules. Furthermore, benzoylbenzoyl ATP also produced shedding of CD23 and L-selectin with the same agonist EC50 values for both (10 μmol/L). Inactivation of the P2Z/P2X7 receptor by preincubation with oxidized ATP abolished ATP-induced shedding of both molecules. Moreover, KN-62, the most potent inhibitor for the P2Z/P2X7 receptor, inhibited ATP-induced shedding of both CD23 and L-selectin with the same IC50 (12 nmol/L). Ro 31-9790, a membrane permeant zinc chelator that inhibits the phorbol-ester-stimulated shedding of L-selectin, also inhibited shedding of CD23 from B-CLL lymphocytes. However, the IC50 for this inhibition by Ro31-9790 was different for L-selectin and CD23 (83 v 6 μmol/L, respectively). Although L-selectin was completely shed by incubation of cells with phorbol-ester, CD23 was not lost under these conditions. The data show that extracellular ATP induces shedding of L-selectin and CD23 from B-CLL lymphocytes by an action mediated by the P2Z/P2X7 receptor. However, different membrane metalloproteases seem to mediate the shedding of L-selectin and CD23. © 1998 by The American Society of Hematology.

Functional expression of purinergic P2X7 receptors in pregnant rat myometrium

2010 ◽  
Vol 298 (4) ◽  
pp. R1117-R1124 ◽  
Author(s):  
Hiroshi Miyoshi ◽  
Kaoru Yamaoka ◽  
Satoshi Urabe ◽  
Miho Kodama ◽  
Yoshiki Kudo
Keyword(s):  
P2x7 Receptor ◽  
Uterine Contraction ◽  
Extracellular Atp ◽  
Receptor Expression ◽  
P2x Receptor ◽  
Receptor Subtype ◽  
P2x7 Receptors ◽  
Pregnant Rat ◽  
Myometrial Cells ◽  
Induced Currents

ATP has been reported to enhance the membrane conductance of myometrial cells and uterine contractility. Purinergic P2 receptor expression has been reported in the myometrium, using molecular biology, but the functional identity of the receptor subtype has not been determined. In this study, ATP-induced currents were recorded and characterized in single myometrial cells from pregnant rats using whole cell patch clamping. Extracellular ATP was applied in the range of 10 μM-1 mM and induced currents with an EC50 of 74 μM, with no desensitization, time dependency, or voltage dependency. The currents induced carried multiple monovalent cations, with conductances ranked as K+ > Cs+ > Li+ > Na+. They were activated by P2X receptor agonists, with their effectiveness ranked as 2′,3′- O-(4-benzoylbenzoyl)-ATP >> ATP > αβ-methylene-ATP > 2-methylthio ATP ≥ UTP ≥ GTP > ADP. These currents were blocked by the selective P2X7 receptor antagonist 3-[5-(2,3-dichlorophenyl)-1 H-tetrazol-1-yl]methyl pyridine (A-438079). We therefore concluded that ATP-induced currents in rat myometrial cells crossed cell membranes via P2X7 receptors. We further showed that the ATP-induced currents were blocked by extracellular Mg2+ (IC50 = 0.26 mM). Clinically, administering extracellular Mg2+ is known to inhibit uterine contraction. It therefore seems likely that uterine contraction may be induced by raised extracellular ATP and suppressed via Mg2+ inhibiting P2X7 receptors. Further research is needed into the P2X7 receptor as a therapeutic target in abnormal uterine contraction, as a possible treatment for premature labor.

scholarly journals P2X7 receptor activates extracellular signal-regulated kinases ERK1 and ERK2 independently of Ca2+ influx

2003 ◽  
Vol 374 (1) ◽  
pp. 51-61 ◽  
Author(s):  
Jan AMSTRUP ◽  
Ivana NOVAK
Keyword(s):  
P2x7 Receptor ◽  
Fluorescent Protein ◽  
Expression System ◽  
Receptor Expression ◽  
Nucleotide Receptors ◽  
P2x7 Receptors ◽  
Hek 293 ◽  
Hek 293 Cells ◽  
293 Cells ◽  
Extracellular Signal

P2X7 nucleotide receptors modulate a spectrum of cellular events in various cells including epithelia, such as exocrine pancreas. Although the pharmacology and channel properties of the P2X7 receptors have been studied intensively, signal transduction pathways are relatively unknown. In this study we applied a heterologous expression system of rat P2X7 receptors in HEK-293 cells. We followed the receptor expression and function using the enhanced green fluorescent protein (EGFP) tag, activation of intracellular proteins and increases in cellular Ca2+. EGFP-P2X7 receptors localized to the plasma membrane, clusters within the membrane and intracellularly. Stimulation of P2X7 receptors in HEK-293 cells led to an activation of extracellular signal-regulated kinases ERK1 and ERK2 and this activation was seen after just 1 min of stimulation with ATP. Using C- and N-terminal P2X7-receptor mutants we show that the N-terminus is important in activation of ERKs, whereas deletion of the last 230 amino acids in the C-terminus did not effect ERK activation. On the other hand, Ca2+ entry was impaired in C-terminal but not in N-terminal mutants. In cell suspensions prepared from rat pancreas we show that P2X7 receptors also activate ERK1 and ERK2, indicating that these signalling pathways are also turned on in native epithelium.

scholarly journals Length of the Neurogenic Period—A Key Determinant for the Generation of Upper-Layer Neurons During Neocortex Development and Evolution

2021 ◽  
Vol 9 ◽  
Author(s):  
Barbara K. Stepien ◽  
Samir Vaid ◽  
Wieland B. Huttner
Keyword(s):  
Cognitive Abilities ◽  
Cortical Neurons ◽  
Mammalian Species ◽  
Critical Role ◽  
Exact Nature ◽  
Cortical Neurogenesis ◽  
Brain Functions ◽  
The Relationship ◽  
Development And Evolution

The neocortex, a six-layer neuronal brain structure that arose during the evolution of, and is unique to, mammals, is the seat of higher order brain functions responsible for human cognitive abilities. Despite its recent evolutionary origin, it shows a striking variability in size and folding complexity even among closely related mammalian species. In most mammals, cortical neurogenesis occurs prenatally, and its length correlates with the length of gestation. The evolutionary expansion of the neocortex, notably in human, is associated with an increase in the number of neurons, particularly within its upper layers. Various mechanisms have been proposed and investigated to explain the evolutionary enlargement of the human neocortex, focussing in particular on changes pertaining to neural progenitor types and their division modes, driven in part by the emergence of human-specific genes with novel functions. These led to an amplification of the progenitor pool size, which affects the rate and timing of neuron production. In addition, in early theoretical studies, another mechanism of neocortex expansion was proposed—the lengthening of the neurogenic period. A critical role of neurogenic period length in determining neocortical neuron number was subsequently supported by mathematical modeling studies. Recently, we have provided experimental evidence in rodents directly supporting the mechanism of extending neurogenesis to specifically increase the number of upper-layer cortical neurons. Moreover, our study examined the relationship between cortical neurogenesis and gestation, linking the extension of the neurogenic period to the maternal environment. As the exact nature of factors promoting neurogenic period prolongation, as well as the generalization of this mechanism for evolutionary distinct lineages, remain elusive, the directions for future studies are outlined and discussed.

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