scholarly journals Cas9-mediated Genome Editing Reveals a Significant Contribution of Calcium Signaling Pathways to Anhydrobiosis in Pv11

2020 ◽  
Author(s):  
Yugo Miyata ◽  
Hiroto Fuse ◽  
Shoko Tokumoto ◽  
Yusuke Hiki ◽  
Ruslan Deviatiiarov ◽  
...  
Keyword(s):  
Calcium Signaling ◽  
Signaling Pathways ◽  
Genome Editing ◽  
Desiccation Tolerance ◽  
Significant Contribution ◽  
Insect Cell Line ◽  
Calmodulin Kinase ◽  
Intracellular Ca ◽  
Polypedilum Vanderplanki ◽  
Creb Pathway

AbstractPv11 is an insect cell line established from the midge Polypedilum vanderplanki that exhibits an extreme desiccation tolerance known as anhydrobiosis. Pv11 has also an anhydrobiotic ability which is induced by trehalose treatment. Here we report the successful construction of the genome editing system for Pv11 cells and its application for identifying the signaling pathways in the anhydrobiosis. Using the Cas9-mediated gene knock-in system, we established GCaMP3-stably expressing Pv11 cells to monitor intracellular Ca2+ mobilization. Intriguingly, trehalose treatment evoked a transient increase of cytosolic Ca2+ concentration, and further experiments indicated the contribution of the calmodulin – calcineurin – NFAT pathway to the tolerance for trehalose treatment as well as the desiccation tolerance, while the calmodulin – calmodulin Kinase – CREB pathway conferred only the desiccation tolerance on Pv11 cells. Thus, our results show the critical contribution of the trehalose–induced Ca2+ surge to the anhydrobiosis and the temporal different roles of each signaling pathway.

scholarly journals Cas9-mediated genome editing reveals a significant contribution of calcium signaling pathways to anhydrobiosis in Pv11 cells

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Yugo Miyata ◽  
Hiroto Fuse ◽  
Shoko Tokumoto ◽  
Yusuke Hiki ◽  
Ruslan Deviatiiarov ◽  
...  
Keyword(s):  
Signaling Pathways ◽  
Genome Editing ◽  
Desiccation Tolerance ◽  
Significant Contribution ◽  
Insect Cell Line ◽  
Calmodulin Kinase ◽  
A Genome ◽  
Intracellular Ca ◽  
Polypedilum Vanderplanki ◽  
Creb Pathway

AbstractPv11 is an insect cell line established from the midge Polypedilum vanderplanki, whose larval form exhibits an extreme desiccation tolerance known as anhydrobiosis. Pv11 itself is also capable of anhydrobiosis, which is induced by trehalose treatment. Here we report the successful construction of a genome editing system for Pv11 cells and its application to the identification of signaling pathways involved in anhydrobiosis. Using the Cas9-mediated gene knock-in system, we established Pv11 cells that stably expressed GCaMP3 to monitor intracellular Ca2+ mobilization. Intriguingly, trehalose treatment evoked a transient increase in cytosolic Ca2+ concentration, and further experiments revealed that the calmodulin–calcineurin–NFAT pathway contributes to tolerance of trehalose treatment as well as desiccation tolerance, while the calmodulin–calmodulin kinase–CREB pathway conferred only desiccation tolerance on Pv11 cells. Thus, our results show a critical contribution of the trehalose-induced Ca2+ surge to anhydrobiosis and demonstrate temporally different roles for each signaling pathway.

Norepinephrine-induced calcium signaling and expression of adrenoceptors in avian tendon cells

2004 ◽  
Vol 287 (4) ◽  
pp. C912-C918 ◽  
Author(s):  
Michelle E. Wall ◽  
James E. Faber ◽  
Xi Yang ◽  
Mari Tsuzaki ◽  
Albert J. Banes
Keyword(s):  
Calcium Signaling ◽  
Mrna Expression ◽  
Signaling Pathways ◽  
Cell Types ◽  
Rt Pcr ◽  
Fura 2 ◽  
Adrenoceptor Antagonist ◽  
Tendon Cells ◽  
Intracellular Ca

Sympathetic efferent nerves are present in tendons, but their function within tendon is unknown. α1-Adrenoceptors are expressed by a variety of cell types. In the presence of norepinephrine (NE), adrenoceptors activate Gq/11 signaling pathways that subsequently increase intracellular Ca2+ concentration ([Ca2+]ic). It was hypothesized that avian tendon cells express functional adrenoceptors that respond to NE by increasing [Ca2+]ic. Avian tendon cells were analyzed for mRNA expression of α1-adrenoceptors by RT-PCR. Avian tendons expressed the α1A- and α1B-adrenoceptor subtypes. Furthermore, both tendon surface epitenon cells and internal fibroblasts infused with a Ca2+-sensitive dye, fura 2, and stimulated with NE responded by increasing [Ca2+]ic. KMD-3213, an α1A-adrenoceptor antagonist, significantly reduced the Ca2+ response. Other adrenoceptor antagonists had no effect on the Ca2+ response. The absence of extracellular Ca2+ also significantly reduced the response to NE, indicating that Ca2+ influx contributed to the rise in [Ca2+]ic. This study provides the first evidence that tendon cells express adrenoceptors and that the NE-induced Ca2+ response is coupled to the α1A-adrenoceptor subtype.

scholarly journals Genome-Wide Role of HSF1 in Transcriptional Regulation of Desiccation Tolerance in the Anhydrobiotic Cell Line, Pv11

2021 ◽  
Vol 22 (11) ◽  
pp. 5798
Author(s):  
Shoko Tokumoto ◽  
Yugo Miyata ◽  
Ruslan Deviatiiarov ◽  
Takahiro G. Yamada ◽  
Yusuke Hiki ◽  
...  
Keyword(s):  
Cell Line ◽  
Cell Lines ◽  
Desiccation Tolerance ◽  
Expression Profiles ◽  
Insect Cell Line ◽  
Gene Expression Profiles ◽  
Late Embryogenesis Abundant ◽  
Heat Shock Factor 1 ◽  
Genome Wide ◽  
A Genome

The Pv11, an insect cell line established from the midge Polypedilum vanderplanki, is capable of extreme hypometabolic desiccation tolerance, so-called anhydrobiosis. We previously discovered that heat shock factor 1 (HSF1) contributes to the acquisition of desiccation tolerance by Pv11 cells, but the mechanistic details have yet to be elucidated. Here, by analyzing the gene expression profiles of newly established HSF1-knockout and -rescue cell lines, we show that HSF1 has a genome-wide effect on gene regulation in Pv11. The HSF1-knockout cells exhibit a reduced desiccation survival rate, but this is completely restored in HSF1-rescue cells. By comparing mRNA profiles of the two cell lines, we reveal that HSF1 induces anhydrobiosis-related genes, especially genes encoding late embryogenesis abundant proteins and thioredoxins, but represses a group of genes involved in basal cellular processes, thus promoting an extreme hypometabolism state in the cell. In addition, HSF1 binding motifs are enriched in the promoters of anhydrobiosis-related genes and we demonstrate binding of HSF1 to these promoters by ChIP-qPCR. Thus, HSF1 directly regulates the transcription of anhydrobiosis-related genes and consequently plays a pivotal role in the induction of anhydrobiotic ability in Pv11 cells.

Involvement of protein kinase C, tyrosine kinases, and Rho kinase in Ca2+ handling of human small arteries

2000 ◽  
Vol 279 (3) ◽  
pp. H1228-H1238 ◽  
Author(s):  
M. Carmen Martínez ◽  
Voahanginirina Randriamboavonjy ◽  
Patrick Ohlmann ◽  
Narcisse Komas ◽  
Juan Duarte ◽  
...  
Keyword(s):  
Protein Kinase C ◽  
Protein Kinase ◽  
Signaling Pathways ◽  
Tyrosine Kinases ◽  
Rho Kinase ◽  
Free Medium ◽  
Kinase C ◽  
Intracellular Ca ◽  
Pkc Inhibitors ◽  
Continuous Presence

The mechanisms of Ca2+ handling and sensitization were investigated in human small omental arteries exposed to norepinephrine (NE) and to the thromboxane A2 analog U-46619. Contractions elicited by NE and U-46619 were associated with an increase in intracellular Ca2+ concentration ([Ca2+]i), an increase in Ca2+-independent signaling pathways, or an enhancement of the sensitivity of the myofilaments to Ca2+. The two latter pathways were abolished by protein kinase C (PKC), tyrosine kinase (TK), and Rho-associated protein kinase (ROK) inhibitors. In Ca2+-free medium, both NE and U-46619 elicited an increase in tension that was greatly reduced by PKC inhibitors and abolished by caffeine or ryanodine. After depletion of Ca2+ stores with NE and U-46619 in Ca2+-free medium, addition of CaCl2 in the continuous presence of the agonists produced increases in [Ca2+]i and contractions that were inhibited by nitrendipine and TK inhibitors but not affected by PKC inhibitors. NE and U-46619 induced tyrosine phosphorylation of a 42- or a 58-kDa protein, respectively. These results indicate that the mechanisms leading to contraction elicited by NE and U-46619 in human small omental arteries are composed of Ca2+ release from ryanodine-sensitive stores, Ca2+ influx through nitrendipine-sensitive channels, and Ca2+ sensitization and/or Ca2+-independent pathways. They also show that the TK pathway is involved in the tonic contraction associated with Ca2+ entry, whereas TK, PKC, and ROK mechanisms regulate Ca2+-independent signaling pathways or Ca2+sensitization.

FEATURES OF CALCIUM HOMEOSTASIS AND CALCIUM SIGNALING IN NEURONS

Vestnik ◽  
2021 ◽  
pp. 208-214
Author(s):  
Б.К. Кайрат ◽  
С.Т. Тулеуханов ◽  
В.П. Зинченко
Keyword(s):  
Plasma Membrane ◽  
Calcium Signaling ◽  
Calcium Binding ◽  
Intracellular Signaling ◽  
Cell Damage ◽  
Compensatory Mechanisms ◽  
Physiological Processes ◽  
Intracellular Ca ◽  
Ca Ions ◽  
Ca Homeostasis

Ионы Са являются основным мессенджером в регуляции физиологических функций клеток. Внутриклеточном пространстве ионы Ca могут свободно состоянии диффундироваться в различных частях цитоплазмы, в то же время значительное количество Ca в связанном виде накапливается в различных внутриклеточных депо или в составе кальций-связывающих белков. Регуляция физиологических процессов с ионами внутриклеточного Са происходит в диапазоне концентраций 10 М, тогда как концентрация Са во внеклеточном пространстве выше и составляет 10 М, для поддержании градиента концентраций в клетках имеются важные Са транспортирующие системы плазматической мембраны, эндоплазматического ретикулума и митохондрий. В нейронах функционируют внутриклеточные ферменты и белки плазматической мембраны для поддержания Са-гомеостаза и реализации механизмов внутриклеточной сигнализации для обеспечения жизнедеятельности в выживании клеток. Нарушение или гиперактивация одного или нескольких механизмов кальциевой сигнализации может привести к повреждению и гибели нейронов в случае отсутствия компенсаторных механизмов. Ca ions are a key messenger for the regulation of most of the physiological functions of cells. Inside the cell, Ca ions can freely diffuse in various parts of the cytoplasm, but a significant amount of Ca is also bound in various intracellular depots or in the form of calcium-binding proteins. The regulation of physiological processes by intracellular Ca ions occurs in the concentration range of 10 M, and the concentration of Ca in the extracellular space is higher and is 10 M, and to maintain this concentration gradient, cells have Ca-transporting systems of the plasma membrane, endoplasmic reticulum and mitochondria. In neurons, a large number of intracellular enzymes and plasma membrane proteins function to maintain Ca-homeostasis and implement intracellular signaling mechanisms to ensure vital activity in the survival of cells. Violation or hyperactivation of one or more mechanisms of calcium signaling can lead to cell damage and death in the absence of compensatory mechanisms.

Bombesin-evoked gastrin release and calcium signaling in human antral G cells in culture

1999 ◽  
Vol 276 (1) ◽  
pp. G227-G237 ◽  
Author(s):  
Paul E. Squires ◽  
R. Mark Meloche ◽  
Alison M. J. Buchan
Keyword(s):  
Cell Culture ◽  
Calcium Signaling ◽  
Gastrin Release ◽  
Voltage Gated ◽  
Gastrin Releasing Peptide ◽  
G Cells ◽  
Peptide Family ◽  
Inositol 1,4,5 Trisphosphate ◽  
Voltage Dependent ◽  
Intracellular Ca

Amplification of mRNA from a human antral cell culture preparation demonstrated the presence of two receptors of the bombesin and gastrin-releasing peptide family, GRPR-1 and BRS-3. Single cell microfluorometry demonstrated that most cells that exhibited bombesin-evoked changes in intracellular Ca2+ concentration were gastrin immunoreactive, indicating that antral G cells express the GRPR subtype. There were two components to the intracellular Ca2+ response: an initial nitrendipine-insensitive mobilization followed by a sustained phase that was inhibited by removal of extracellular Ca2+ and 20 mM caffeine and was partially inhibited by 10 μM nitrendipine. Preexposure of cells to thapsigargin and caffeine prevented the response to bombesin, indicating activation of inositol 1,4,5-trisphosphate (IP3)-sensitive stores. Gastrin release could be partially reversed by removal of extracellular Ca2+ and blockade of L-type voltage-dependent Ca2+ channels, indicating that a component of the secretory response to bombesin was dependent on Ca2+ influx. These data demonstrated that bombesin-stimulated gastrin release from human antral G cells resulted from activation of GRPRs and involved both release of intracellular Ca2+ and influx of extracellular Ca2+through a combination of L-type voltage-gated and IP3-gated Ca2+ channels.

scholarly journals Ozone-Induced Cell Death Mediated with Oxidative and Calcium Signaling Pathways in Tobacco Bel-W3 and Bel-B Cell Suspension Cultures

2006 ◽  
Vol 1 (6) ◽  
pp. 312-322 ◽  
Author(s):  
Takashi Kadono ◽  
Yuka Yamaguchi ◽  
Takuya Furuichi ◽  
Manabu Hirono ◽  
Jean-Pierre Garrec ◽  
...  
Keyword(s):  
Cell Death ◽  
Calcium Signaling ◽  
B Cell ◽  
Cell Suspension ◽  
Signaling Pathways ◽  
Cell Suspension Cultures ◽  
Suspension Cultures

Glucose is a key metabolic regulator of osteoclasts; glucose stimulated increases in ATP/ADP ratio and calmodulin kinase II activity

2005 ◽  
Vol 83 (5) ◽  
pp. 667-673 ◽  
Author(s):  
Kirsten I Larsen ◽  
Marina Falany ◽  
Wei Wang ◽  
John P Williams
Keyword(s):  
Glucose Transport ◽  
Glucose Sensing ◽  
Mitogen Activated Protein Kinase ◽  
Β Cells ◽  
Calcium Calmodulin Dependent ◽  
Calmodulin Kinase ◽  
Rapid Changes ◽  
Mitogen Activated Protein ◽  
Intracellular Ca ◽  
Dependent Signaling Pathway

Glucose-stimulated increases in osteoclast activity are mediated, at least in part, by transcriptional regulation of H+-ATPase expression through a mechanism involving p38 mitogen-activated protein kinase. We hypothesized that early events in the glucose-dependent signaling pathway would be similar to those identified in other glucose-sensitive cells, such as islet β-cells, including rapid changes in the cellular ATP/ADP ratio and mobilization of intracellular Ca2+. We demonstrate that glucose stimulates a prolonged 50% increase in the ATP/ADP ratio that was maximal 30 s after glucose concentrations were increased. Glucose stimulated a transient 30% increase in calcium/calmodulin-dependent kinase II (CaMK II) activity that was maximal 3 min after the glucose concentration was increased. CaMK II was activated maximally by 3 mmol D-glucose/L in 3-min assays. Activation of CaMK II in the presence of the nonmetabo lizable glucose analog 2-deoxyglucose was 2-fold greater than with D-glucose but was unchanged by glucosamine. Pretreatment of osteoclasts with the intracellular Ca2+ chelator BAPTA-AM inhibited glucose transport by 75%. BAPTA-AM treatment also prevented glucose-dependent stimulation of CaMK II. The data indicate that osteoclasts utilize a glucose-sensing mechanism similar to that of β-cells and that glucose-stimulated signaling in osteoclasts involves changes in the ATP/ADP ratio and mobilization of intracellular Ca2+, resulting in activation of CaMK II.Key words: osteoclast, metabolism, glucose transport, calmodulin kinase II.

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