scholarly journals Melatonin promotes sleep by activating the BK channel in C. elegans

2020 ◽  
Vol 117 (40) ◽  
pp. 25128-25137
Author(s):  
Longgang Niu ◽  
Yan Li ◽  
Pengyu Zong ◽  
Ping Liu ◽  
Yuan Shui ◽  
...  
Keyword(s):  
G Protein ◽  
Neurotransmitter Release ◽  
Expression System ◽  
Molecular Target ◽  
Bk Channel ◽  
G Protein Coupled Receptors ◽  
Wild Type ◽  
Heterologous Expression System ◽  
C Elegans ◽  
G Protein Coupled

Melatonin (Mel) promotes sleep through G protein-coupled receptors. However, the downstream molecular target(s) is unknown. We identified the Caenorhabditis elegans BK channel SLO-1 as a molecular target of the Mel receptor PCDR-1-. Knockout of pcdr-1, slo-1, or homt-1 (a gene required for Mel synthesis) causes substantially increased neurotransmitter release and shortened sleep duration, and these effects are nonadditive in double knockouts. Exogenous Mel inhibits neurotransmitter release and promotes sleep in wild-type (WT) but not pcdr-1 and slo-1 mutants. In a heterologous expression system, Mel activates the human BK channel (hSlo1) in a membrane-delimited manner in the presence of the Mel receptor MT1 but not MT2. A peptide acting to release free Gβγ also activates hSlo1 in a MT1-dependent and membrane-delimited manner, whereas a Gβλ inhibitor abolishes the stimulating effect of Mel. Our results suggest that Mel promotes sleep by activating the BK channel through a specific Mel receptor and Gβλ.

scholarly journals Rescue of defective G protein–coupled receptor function in vivo by intermolecular cooperation

2010 ◽  
Vol 107 (5) ◽  
pp. 2319-2324 ◽  
Author(s):  
Adolfo Rivero-Müller ◽  
Yen-Yin Chou ◽  
Inhae Ji ◽  
Svetlana Lajic ◽  
Aylin C. Hanyaloglu ◽  
...  
Keyword(s):  
G Protein ◽  
G Protein Coupled Receptors ◽  
Luteinizing Hormone Receptor ◽  
Wild Type ◽  
Gpcr Signaling ◽  
Physiological Relevance ◽  
Biosynthesis Activation ◽  
Mutant Receptors ◽  
G Protein Coupled

G protein–coupled receptors (GPCRs) are ubiquitous mediators of signaling of hormones, neurotransmitters, and sensing. The old dogma is that a one ligand/one receptor complex constitutes the functional unit of GPCR signaling. However, there is mounting evidence that some GPCRs form dimers or oligomers during their biosynthesis, activation, inactivation, and/or internalization. This evidence has been obtained exclusively from cell culture experiments, and proof for the physiological significance of GPCR di/oligomerization in vivo is still missing. Using the mouse luteinizing hormone receptor (LHR) as a model GPCR, we demonstrate that transgenic mice coexpressing binding-deficient and signaling-deficient forms of LHR can reestablish normal LH actions through intermolecular functional complementation of the mutant receptors in the absence of functional wild-type receptors. These results provide compelling in vivo evidence for the physiological relevance of intermolecular cooperation in GPCR signaling.

scholarly journals Selective Modulation of Wild Type Receptor Functions by Mutants of G-Protein-coupled Receptors

1999 ◽  
Vol 274 (18) ◽  
pp. 12548-12554 ◽  
Author(s):  
Christian Le Gouill ◽  
Jean-Luc Parent ◽  
Carolyn-Ann Caron ◽  
Rémi Gaudreau ◽  
Léonid Volkov ◽  
...  
Keyword(s):  
G Protein ◽  
G Protein Coupled Receptors ◽  
Wild Type ◽  
Selective Modulation ◽  
G Protein Coupled ◽  
Type Receptor

scholarly journals Identification of a Conserved, Orphan G Protein-Coupled Receptor Required for Efficient Pathogen Clearance in Caenorhabditis elegans

2019 ◽  
Vol 87 (4) ◽  
Author(s):  
Alexandra Anderson ◽  
Yee Lian Chew ◽  
William Schafer ◽  
Rachel McMullan
Keyword(s):  
Caenorhabditis Elegans ◽  
Immune Responses ◽  
G Protein ◽  
G Protein Coupled Receptors ◽  
G Protein Coupled Receptor ◽  
Content Type ◽  
C Elegans ◽  
Host Immune Responses ◽  
Pathogen Clearance ◽  
G Protein Coupled

ABSTRACT G protein-coupled receptors contribute to host defense across the animal kingdom, transducing many signals involved in both vertebrate and invertebrate immune responses. While it has become well established that the nematode worm Caenorhabditis elegans triggers innate immune responses following infection with numerous bacterial, fungal, and viral pathogens, the mechanisms by which C. elegans recognizes these pathogens have remained somewhat more elusive. C. elegans G protein-coupled receptors have been implicated in recognizing pathogen-associated damage and activating downstream host immune responses. Here we identify and characterize a novel G protein-coupled receptor required to regulate the C. elegans response to infection with Microbacterium nematophilum. We show that this receptor, which we designate pathogen clearance-defective receptor 1 (PCDR-1), is required for efficient pathogen clearance following infection. PCDR-1 acts upstream of multiple G proteins, including the C. elegans Gαq ortholog, EGL-30, in rectal epithelial cells to promote pathogen clearance via a novel mechanism.

High-Level Expression of G Protein-Coupled Receptors with the Aid of the Semliki Forest Virus Expression System

1995 ◽  
Vol 15 (1-4) ◽  
pp. 23-32 ◽  
Author(s):  
Kenneth Lundstrom ◽  
Ann Mills ◽  
Elisabeth Allet ◽  
Karin Ceszkowski ◽  
Georges Agudo ◽  
...  
Keyword(s):  
G Protein ◽  
Semliki Forest Virus ◽  
Expression System ◽  
G Protein Coupled Receptors ◽  
High Level Expression ◽  
Virus Expression ◽  
High Level ◽  
G Protein Coupled

Employing Rhodobacter sphaeroides to functionally express and purify human G protein-coupled receptors

2008 ◽  
Vol 389 (1) ◽  
pp. 69-78 ◽  
Author(s):  
Ankita Roy ◽  
Arun Kumar Shukla ◽  
Winfried Haase ◽  
Hartmut Michel
Keyword(s):  
G Protein ◽  
Rhodobacter Sphaeroides ◽  
Structural Information ◽  
Membrane Surface ◽  
Expression System ◽  
Gene Promoter ◽  
Photosynthetic Bacterium ◽  
G Protein Coupled Receptors ◽  
Recombinant Receptors ◽  
G Protein Coupled

AbstractG protein-coupled receptors (GPCRs) represent the largest class of cell surface receptors and play crucial roles in many cellular and physiological processes. Functional production of recombinant GPCRs is one of the main bottlenecks to obtaining structural information. Here, we report the use of a novel bacterial expression system based on the photosynthetic bacteriumRhodobacter sphaeroidesfor the production of human recombinant GPCRs. The advantage of employingR. sphaeroidesas a host lies in the fact that it provides much more membrane surface per cell compared to other typical expression hosts. The system was tailored to overexpress recombinant receptors under the control of the moderately strong and highly regulated superoperonic photosynthetic promoterpufQ. We tested this system for the expression of some class A GPCRs, namely, the human adenosine A2a receptor (A2aR), the human angiotensin AT1a receptor (AT1aR) and the human bradykinin B2 receptor (B2R). Several different constructs were examined and functional production of the recombinant receptors was achieved. The best-expressed receptor, AT1aR, was solubilized and affinity-purified. To the best of our knowledge, this is the first report of successful use of a bacterial host –R. sphaeroides– to produce functional recombinant GPCRs under the control of a photosynthetic gene promoter.

Caspase-12: a developmental link between G-protein–coupled receptors and integrin αIIbβ3 activation

Blood ◽  
2004 ◽  
Vol 104 (5) ◽  
pp. 1327-1334 ◽  
Author(s):  
Steven W. Kerrigan ◽  
Meenakshi Gaur ◽  
Ronan P. Murphy ◽  
Sanford J. Shattil ◽  
Andrew D. Leavitt
Keyword(s):  
G Protein ◽  
Adenosine Diphosphate ◽  
G Protein Coupled Receptors ◽  
Free Calcium ◽  
Wild Type ◽  
Integrin Αiibβ3 ◽  
Caspase 12 ◽  
Fibrinogen Binding ◽  
G Protein Coupled ◽  
Inside Out

Abstract Fibrinogen binding by integrin αIIbβ3 is promoted by platelet agonists that increase the affinity and avidity of αIIbβ3 for fibrinogen through a process called “inside-out” signaling. Having previously demonstrated that inside-out activation of αIIbβ3 is defective in murine megakaryocytes that lack the transcription factor NF-E2, we screened for NF-E2–regulated genes that affect αIIbβ3 activation. Caspase-12 is the most down-regulated gene we identified in NF-E2–/– megakaryocytes. Therefore, the role of this protein in αIIbβ3 activation was determined using platelets from caspase-12–/– mice. Despite wild-type levels of αIIbβ3, caspase-12–/– platelets exhibit reduced fibrinogen binding to αIIbβ3 following stimulation by adenosine diphosphate (ADP) or protease-activated receptor 4 (PAR4) receptor-activating peptide. The defect in αIIbβ3 activation is associated with decreased cytosolic free calcium and inositol triphosphate levels, and with reduced aggregation, despite wild-type phospholipase Cβ expression levels. In contrast, agonist-induced surface expression of P-selectin, suppression of cAMP levels following ADP stimulation, and spreading on immobilized fibrinogen are unimpaired. Moreover, although caspase-12 is highly expressed in mature megakaryocytes, it is undetectable in platelets. Taken together, these studies establish that caspase-12 expression in murine megakaryocytes is regulated, directly or indirectly, by NF-E2, and suggest that caspase-12 participates in the development of fully functional signaling pathways linking some G-protein–coupled receptors to αIIbβ3 activation.

scholarly journals Structural and functional analysis of the canine histamine H2 receptor by site-directed mutagenesis: N-glycosylation is not vital for its action

1995 ◽  
Vol 310 (2) ◽  
pp. 553-558 ◽  
Author(s):  
Y Fukushima ◽  
Y Oka ◽  
T Saitoh ◽  
H Katagiri ◽  
T Asano ◽  
...  
Keyword(s):  
Molecular Mass ◽  
G Protein ◽  
G Protein Coupled Receptors ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Wild Type ◽  
Histamine H2 Receptor ◽  
H2 Receptor ◽  
H2 Receptors ◽  
G Protein Coupled

G-protein-coupled receptors generally share a similar structure containing seven membrane-spanning domains and extracellular site(s) for N-glycosylation. The histamine H2 receptor is a member of the family of G-protein-coupled receptors, and has three extracellular potential sites for N-glycosylation (Asn-4, Asn-162 and Asn-168). To date, however, no information has been presented regarding N-glycosylation of the H2 receptor. To investigate the presence, location and functional roles of N-glycosylation of the H2 receptor, site-directed mutagenesis was performed to eliminate the potential site(s) for N-glycosylation singly and collectively. The wild-type and mutated H2 receptors were expressed stably in Chinese hamster ovary (CHO) cells or transiently in COS7 cells. Immunoblotting of the wild-type and mutated H2 receptors with an antiserum directed against the C-terminus of the H2 receptor showed that mutation at Asn-162, but not at Asn-168, resulted in a substantial decrease in the molecular mass. A mutation at Asn-4 led to a further decrease in the molecular mass. Tunicamycin treatment of the transfected cells yielded a sharp band with a molecular mass identical to that of the mutant devoid of all three potential sites for N-glycosylation. These findings indicate that the H2 receptor is N-glycosylated, and that N-glycosylation takes place mainly at two sites, Asn-4 and Asn-162. Neither the affinity for tiotidine nor that for histamine was affected by the mutagenesis. Immunocytochemistry and tiotidine binding showed that the mutated receptors were exclusively distributed on the cell surface in a fashion similar to that of the wild-type. In addition, the glycosylation-defective receptor was capable of activating adenylate cyclase and elevating the intracellular Ca2+ concentration in response to histamine in stable CHO cell lines. Thus N-glycosylation of the H2 receptor is not required for cell surface localization, ligand binding or functional coupling to G-protein(s).

Sign in / Sign up

Export Citation Format

Share Document