scholarly journals A75 ON THE TRAIL OF ALTERNATIVE LIGANDS AND SIGNALING PATHWAYS FOR LGR5

2020 ◽  
Vol 3 (Supplement_1) ◽  
pp. 89-90
Author(s):  
S Dagenais Bellefeuille ◽  
G Arguin ◽  
F Gendron
Keyword(s):  
Signaling Network ◽  
Bioactive Molecules ◽  
Orphan Receptor ◽  
Hek293 Cells ◽  
Intestinal Lumen ◽  
Aberrant Expression ◽  
Alternative Pathways ◽  
Canonical Wnt ◽  
Complex Signaling ◽  
Stimulation Of

Abstract Background Leucine-rich G protein coupled receptor-5 (LGR5) is a GPCR originally identified as a marker for intestinal stem cells but is now associated to stemness in numerous other tissues. R-spondins (RSPO) are the only reported ligands of LGR5, which upon receptor binding, potentiates the canonical Wnt/b-catenin pathway. Surprisingly, despite the presence of classical GPCR features such as conserved DRY and NPXXY motifs RSPO binding to LGR5 does not induce classical GPCR behaviors such as coupling to heterotrimeric G-proteins or engagement of b-arrestins. For this reason, LGR5 is still considered to be an orphan receptor. Aims Aberrant expression of LGR5 is a common feature of many cancers, including gastrointestinal cancers, but it is still unclear why LGR5 is pro-tumorigenic in some cancers, while anti-tumorigenic in others. One potential explanation is that LGR5 does not only signals through the Wnt/b-catenin pathway, but also to alternative pathways. Thus, the goals of this study were 1), to elucidate LGR5 signaling networks and 2), to identify new LGR5 ligands and activity modulators. We suggested that the intestinal lumen content could be a source of bioactive molecules that could act as alternative LGR5 ligands Methods All experiments were performed on HEK293 cells expressing recombinant LGR5 (HEK293/LGR5) and compared to control cells expressing empty vector. Characterization of LGR5 signaling network was initiated with LC MS/MS spectrometric analyses using cells stimulated with Wnt3a and RSPO1. To identify new LGR5 modulators, commercially available bacteria-derived metabolites (BDM) were tested on HEK293/LGR5 cells stimulated with Wnt3a and RSPO1 and assessed using the TOP-Flash luciferase system. Finally, a mouse intestinal lumen extract (ILE) was investigated in whole-cell impedance assay to detect if it contained native BDM or potential host-derived LGR5 ligands. Results Mass spectrometric analyses revealed distinct protein expression and phosphorylation profiles in HEK293/LGR5 cells stimulated with Wnt3a and RSPO1. The TOP/Flash luciferase assays showed that the flavonoid derivative 3,4-dihydroxyphenylacetic acid (DOPAC) and lipopolysaccharide (LPS) acted as negative regulators of LGR5, while butyrate had no effect. Interestingly, stimulation of HEK293/LGR5 with ILE induced a Gαq-like response in the impedance assay, while control cells did not respond. Conclusions Stimulation of LGR5 with Wnt and RSPO triggered a complex signaling network. In agreement with our hypothesis, some BDM such as DOPAC and LPS were indeed activity modulators of LGR5. Moreover, a still unidentified compound present in the ILE triggered a selective GPCR-like response in HEK293/LGR5 cells. Future works aim at identifying this putative LGR5 ligand(s) and establish the LGR5 interactome using proximity labeling and mass spectrometry. Funding Agencies NSERC, Université de Sherbrooke, FRQS

Role of the Go/i signaling network in the regulation of neurite outgrowthThis paper is one of a selection of papers published in this Special issue, entitled Second Messengers and Phosphoproteins—12th International Conference.

2006 ◽  
Vol 84 (7) ◽  
pp. 687-694 ◽  
Author(s):  
John Cijiang He ◽  
Susana R. Neves ◽  
J. Dedrick Jordan ◽  
Ravi Iyengar
Keyword(s):  
Neurite Outgrowth ◽  
Cannabinoid Receptor ◽  
Second Messengers ◽  
Signaling Network ◽  
Neuronal Growth ◽  
Alternative Pathways ◽  
Cb1 Cannabinoid Receptor ◽  
Neuro2a Cells ◽  
Complex Signaling ◽  
Rap1 Activation

Neurite outgrowth is a complex differentiation process stimulated by many neuronal growth factors and transmitters and by electrical activity. Among these stimuli are ligands for G-protein-coupled receptors (GPCR) that function as neurotransmitters. The pathways involved in GPCR-triggered neurite outgrowth are not fully understood. Many of these receptors couple to Gαo, one of the most abundant proteins in the neuronal growth cones. We have studied the Go signaling network involved in neurite outgrowth in Neuro2A cells. Gαo can induce neurite outgrowth. The CB1 cannabinoid receptor, a Go/i-coupled receptor expressed endogenously in Neuro2A cells, triggers neurite outgrowth by activating Rap1, which promotes the Gαo-stimulated proteasomal degradation of Rap1GAPII. CB1-receptor-mediated Rap1 activation leads to the activation of a signaling network that includes the small guanosine triphosphate (GTP)ases Ral and Rac, the protein kinases Src, and c-Jun N-terminal kinase (JNK), which converge onto the activation of signal transducer and activator of transcription 3 (Stat3), a key transcription factor that mediates the gene expression process of neurite outgrowth in Neuro2A cells. This review describes current findings from our laboratory and also discusses alternative pathways that Go/i might mediate to trigger neurite outgrowth. We also analyze the role neurotransmitters, which stimulate Go/i to activate a complex signaling network controlling neurite outgrowth, play in regeneration after neuronal injury.

scholarly journals Alternative pathways for the development of lymphoid structures in humans

2021 ◽  
Vol 118 (29) ◽  
pp. e2108082118
Author(s):  
Laureline Berteloot ◽  
Thierry Jo Molina ◽  
Julie Bruneau ◽  
Capucine Picard ◽  
Vincent Barlogis ◽  
...  
Keyword(s):  
T Cell ◽  
Severe Combined Immunodeficiency ◽  
Lymphoid Organs ◽  
Lymphoid Cells ◽  
Orphan Receptor ◽  
Alternative Pathways ◽  
Cell Engraftment ◽  
Host Origin ◽  
Corticomedullary Differentiation ◽  
Lymphoid Structures

Lymphoid tissue inducer (LTi) cells are critical for inducing the differentiation of most secondary lymphoid organs (SLOs) in mice. In humans, JAK3 and γc deficiencies result in severe combined immunodeficiency (SCIDs) characterized by an absence of T cells, natural killer cells, innate lymphoid cells (ILCs), and presumably LTi cells. Some of these patients have undergone allogeneic stem cell transplantation (HSCT) in the absence of myeloablation, which leads to donor T cell engraftment, while other leukocyte subsets are of host origin. By using MRI to look for SLOs in nine of these patients 16 to 44 y after HSCT, we discovered that SLOs were exclusively found in the three areas of the abdomen that drain the intestinal tract. A postmortem examination of a child with γc-SCID who had died 3.5 mo after HSCT showed corticomedullary differentiation in the thymus, T cell zones in the spleen, and the appendix, but in neither lymph nodes nor Peyer patches. Tertiary lymphoid organs were observed in the lung. No RAR-related orphan receptor-positive LTi cells could be detected in the existing lymphoid structures. These results suggest that while LTi cells are required for the genesis of most SLOs in humans, SLO in the appendix and in gut-draining areas, as well as tertiary lymphoid organs, can be generated likely by LTi cell-independent mechanisms.

scholarly journals Serotonin 2A (5-HT2A) receptor affects cell–matrix adhesion and the formation and maintenance of stress fibers in HEK293 cells

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Joe Anand Kumar John Jayakumar ◽  
Mitradas. M. Panicker ◽  
Basudha Basu
Keyword(s):  
Nervous System ◽  
Neural Circuits ◽  
Stress Fibers ◽  
Hek293 Cells ◽  
Matrix Adhesion ◽  
Cell Matrix ◽  
Cell Matrix Adhesion ◽  
G Protein Coupled ◽  
Serotonin 2A ◽  
Stimulation Of

Abstract5-HT2A, a G-protein coupled receptor, is widely expressed in the human body, including in the gastrointestinal tract, platelets and the nervous system. It mediates various functions, for e.g. learning, memory, mood regulation, platelet aggregation and vasoconstriction, but its involvement in cell-adhesion remains largely unknown. Here we report a novel role for 5-HT2A in cell–matrix adhesion.In HEK293 cells, which are loosely adherent, expression and stimulation of human or rat 5-HT2A receptor by agonists such as serotonin or 2,5-dimethoxy-4-iodoamphetamine (DOI) led to a significant increase in adhesion, while inhibition of 5-HT2A by antipsychotics, such as risperidone, olanzapine or chlorpromazine prevented it. 5-HT2A activation gave rise to stress fibers in these cells and was also required for their maintenance. Mechanistically, the 5-HT2A-mediated adhesion was mediated by downstream PKC and Rho signaling. Since 5-HT2A is associated with many disorders such as dementia, depression and schizophrenia, its role in cell–matrix adhesion could have implications for neural circuits.

scholarly journals Calmidazolium and arachidonate activate a calcium entry pathway that is distinct from store-operated calcium influx in HeLa cells

2004 ◽  
Vol 381 (3) ◽  
pp. 929-939 ◽  
Author(s):  
Claire M. PEPPIATT ◽  
Anthony M. HOLMES ◽  
Jeong T. SEO ◽  
Martin D. BOOTMAN ◽  
Tony J. COLLINS ◽  
...  
Keyword(s):  
Hela Cells ◽  
Calcium Influx ◽  
Differential Sensitivity ◽  
Excitable Cells ◽  
Hormonal Stimulation ◽  
Calmodulin Antagonist ◽  
Alternative Pathways ◽  
Entry Pathway ◽  
Store Depletion ◽  
Stimulation Of

Agonists that deplete intracellular Ca2+ stores also activate Ca2+ entry, although the mechanism by which store release and Ca2+ influx are linked is unclear. A potential mechanism involves ‘store-operated channels’ that respond to depletion of the intracellular Ca2+ pool. Although SOCE (store-operated Ca2+ entry) has been considered to be the principal route for Ca2+ entry during hormonal stimulation of non-electrically excitable cells, recent evidence has suggested that alternative pathways activated by metabolites such as arachidonic acid are responsible for physiological Ca2+ influx. It is not clear whether such messenger-activated pathways exist in all cells, whether they are truly distinct from SOCE and which metabolites are involved. In the present study, we demonstrate that HeLa cells express two pharmacologically and mechanistically distinct Ca2+ entry pathways. One is the ubiquitous SOCE route and the other is an arachidonate-sensitive non-SOCE. We show that both these Ca2+ entry pathways can provide long-lasting Ca2+ elevations, but that the channels are not the same, based on their differential sensitivity to 2-aminoethoxydiphenyl borate, LOE-908 {(R,S)-(3,4-dihydro-6,7-dimethoxy-isochinolin-1-yl)-2-phenyl-N,N-di[2-(2,3,4-trimethoxyphenyl)ethyl]acetamid mesylate} and gadolinium. In addition, non-SOCE and not SOCE was permeable to strontium. Furthermore, unlike SOCE, the non-SOCE pathway did not require store depletion and was not sensitive to displacement of the endoplasmic reticulum from the plasma membrane using jasplakinolide or ionomycin pretreatment. These pathways did not conduct Ca2+ simultaneously due to the dominant effect of arachidonate, which rapidly curtails SOCE and promotes Ca2+ influx via non-SOCE. Although non-SOCE could be activated by exogenous application of arachidonate, the most robust method for stimulation of this pathway was application of the widely used calmodulin antagonist calmidazolium, due to its ability to activate phospholipase A2.

Alarmin S100A9 Induces Proinflammatory and Catabolic Effects Predominantly in the M1 Macrophages of Human Osteoarthritic Synovium

2016 ◽  
Vol 43 (10) ◽  
pp. 1874-1884 ◽  
Author(s):  
Martijn H. van den Bosch ◽  
Arjen B. Blom ◽  
Rik F. Schelbergen ◽  
Marije I. Koenders ◽  
Fons A. van de Loo ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Proinflammatory Cytokines ◽  
Colony Stimulating Factor ◽  
Canonical Wnt Signaling ◽  
Gm Csf ◽  
Macrophage Colony Stimulating Factor ◽  
Macrophage Colony ◽  
Canonical Wnt ◽  
Stimulating Factor ◽  
Stimulation Of

Objective.The alarmins S100A8 and S100A9 have been shown to regulate synovial activation, cartilage damage, and osteophyte formation in osteoarthritis (OA). Here we investigated the effect of S100A9 on the production of proinflammatory cytokines and matrix metalloprotease (MMP) in OA synovium, granulocyte macrophage colony-stimulating factor (GM-CSF)-differentiated/macrophage colony-stimulating factor (M-CSF)-differentiated macrophages, and OA fibroblasts.Methods.We determined which cell types in the synovium produced S100A8 and S100A9. Further, the production of proinflammatory cytokines and MMP, and the activation of canonical Wnt signaling, was determined in human OA synovium, OA fibroblasts, and monocyte-derived macrophages following stimulation with S100A9.Results.We observed that S100A8 and S100A9 were mainly produced by GM-CSF–differentiated macrophages present in the synovium, and to a lesser extent by M-CSF–differentiated macrophages, but not by fibroblasts. S100A9 stimulation of OA synovial tissue increased the production of the proinflammatory cytokines interleukin (IL) 1β, IL-6, IL-8, and tumor necrosis factor-α. Additionally, various MMP were upregulated after S100A9 stimulation. Experiments to determine which cell type was responsible for these effects revealed that mainly stimulation of GM-CSF–differentiated macrophages and to a lesser extent M-CSF-differentiated macrophages with S100A9 increased the expression of these proinflammatory cytokines and MMP. In contrast, stimulation of fibroblasts with S100A9 did not affect their expression. Finally, stimulation of GM-CSF–differentiated, but not M-CSF–differentiated macrophages with S100A9 activated canonical Wnt signaling, whereas incubation of OA synovium with the S100A9 inhibitor paquinimod reduced the activation of canonical Wnt signaling.Conclusion.Predominantly mediated by M1-like macrophages, the alarmin S100A9 stimulates the production of proinflammatory and catabolic mediators and activates canonical Wnt signaling in OA synovium.

scholarly journals Complex Signaling Network in Regulation of Adenosine 5′-Phosphosulfate Reductase by Salt Stress in Arabidopsis Roots

2008 ◽  
Vol 146 (3) ◽  
pp. 1408-1420 ◽  
Author(s):  
Anna Koprivova ◽  
Kathryn Anne North ◽  
Stanislav Kopriva
Keyword(s):  
Salt Stress ◽  
Signaling Network ◽  
Complex Signaling

Selective sacral rootlet rhizotomy for hypertonic neurogenic bladder

1975 ◽  
Vol 42 (5) ◽  
pp. 567-574 ◽  
Author(s):  
Stanislaw K. Toczek ◽  
David C. McCullough ◽  
Guy W. Gargour ◽  
Rudolf Kachman ◽  
Roger Baker ◽  
...  
Keyword(s):  
Neurogenic Bladder ◽  
Conus Medullaris ◽  
Physiological Data ◽  
Detrusor Muscle ◽  
Lower Extremity Function ◽  
Accurate Identification ◽  
Content Type ◽  
Alternative Pathways ◽  
Bladder Detrusor ◽  
Stimulation Of

✓ The authors describe a highly selective transsacral microsurgical procedure for sacral nerve rootlet interruption in five patients with hypertonic neurogenic bladder. Magnification and systematic stimulation of sacral roots provided accurate identification of motor fibers supplying bladder detrusor muscle and differentiation of efferent components to the legs and anal sphincter. Although the technique prevented incontinence and adverse effects of nerve section on rectal and lower extremity function, improvement in voiding patterns and diminution of urinary sepsis was of brief duration in three of the five patients. Physiological data from these procedures reaffirms the importance of S-3 and S-4 motor roots in detrusor innervation, but clinical responses bring into question the possibility of sustained improvement from such a highly selective procedure at the sacral level. The authors suggest that alternative pathways, not apparent on initial stimulation, may develop after section of sacral root components, and that dissection and stimulation of fibers at the level of the conus medullaris should be investigated as an alternative procedure.

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