scholarly journals Not So Dry After All: DRY Mutants of the AT1A Receptor and H1 Receptor Can Induce G-Protein-Dependent Signaling

ACS Omega ◽  
2020 ◽  
Vol 5 (6) ◽  
pp. 2648-2659
Author(s):  
Anna Pietraszewska-Bogiel ◽  
Linda Joosen ◽  
Anna O. Chertkova ◽  
Joachim Goedhart
Keyword(s):  
G Protein ◽  
H1 Receptor ◽  
At1a Receptor

scholarly journals The Target Residence Time of Antihistamines Determines Their Antagonism of the G Protein-Coupled Histamine H1 Receptor

2017 ◽  
Vol 8 ◽  
Author(s):  
Reggie Bosma ◽  
Gesa Witt ◽  
Lea A. I. Vaas ◽  
Ivana Josimovic ◽  
Philip Gribbon ◽  
...  
Keyword(s):  
G Protein ◽  
Residence Time ◽  
H1 Receptor ◽  
Histamine H1 Receptor ◽  
G Protein Coupled

scholarly journals Not so dry after all – DRY mutants of the AT1A receptor and H1 receptor can induce G protein-dependent signaling

2019 ◽  
Author(s):  
A Pietraszewska-Bogiel ◽  
L Joosen ◽  
J Goedhart
Keyword(s):  
G Protein ◽  
G Proteins ◽  
Intracellular Signaling ◽  
Adaptor Proteins ◽  
Common Belief ◽  
Protein Activation ◽  
Protein Kinase C Activity ◽  
Gpcr Activation ◽  
Successful Isolation ◽  
At1a Receptor

ABSTRACTGPCRs are seven transmembrane spanning receptors that regulate a wide array of intracellular signaling cascades in response to various stimuli. To do so, they couple to different heterotrimeric G proteins and adaptor proteins, including arrestins. Importantly, arrestins were shown to regulate GPCR signaling through G proteins, as well as promote G protein-independent signaling events. Several research groups have reported successful isolation of exclusively G protein-dependent and arrestin-dependent signaling downstream of GPCR activation using biased agonists or receptor mutants incapable of coupling to either arrestins or G proteins. In the latter category, the DRY mutant of the angiotensin II type 1 receptor was extensively used to characterize functional selectivity downstream of AT1AR. In an attempt to understand histamine 1 receptor signaling, we characterized the signaling capacity of the H1R DRY mutant in a panel of dynamic, live cell biosensor assays, including arrestin recruitment, heterotrimeric G-protein activation, Ca2+ signaling, protein kinase C activity, GTP binding of RhoA, and activation of ERK1/2. Here we show that both H1R DRY mutant and the AT1AR DRY mutant (used as a reference) are capable of efficient activation of G protein-mediated signaling. Therefore, contrary to common belief, they do not constitute suitable tools for dissection of arrestin-mediated, G protein-independent signaling downstream of these receptors.

scholarly journals Evidence for involvement of 3'-untranslated region in determining angiotensin II receptor coupling specificity to G-protein

2003 ◽  
Vol 370 (2) ◽  
pp. 631-639 ◽  
Author(s):  
Thomas J. THEKKUMKARA ◽  
Stuart L. LINAS
Keyword(s):  
G Protein ◽  
Untranslated Region ◽  
Chinese Hamster ◽  
Cell Number ◽  
Cross Linking ◽  
Leucine Incorporation ◽  
Wild Type ◽  
Receptor Coupling ◽  
At1a Receptor ◽  
Mrna Binding

The mRNA 3′-untranslated region (3′-UTR) of many genes has been identified as an important regulator of the mRNA transcript itself as well as the translated product. Previously, we demonstrated that Chinese-hamster ovary-K1 cells stably expressing angiotensin receptor subtypes (AT1A) with and without 3′-UTR differed in AT1A mRNA content and its coupling with intracellular signalling pathways. Moreover, RNA mobility-shift assay and UV cross-linking studies using the AT1A 3′-UTR probe identified a major mRNA-binding protein complex of 55kDa in Chinese-hamster ovary-K1 cells. In the present study, we have determined the functional significance of the native AT1A receptor 3′-UTR in rat liver epithelial (WB) cell lines by co-expressing the AT1A 3′-UTR sequence ‘decoy’ to compete with the native receptor 3′-UTR for its mRNA-binding proteins. PCR analysis using specific primers for the AT1A receptor and [125I]angiotensin II (AngII)-binding studies demonstrated the expression of the native AT1A receptors in WB (Bmax = 2.7pmol/mg of protein, Kd = 0.56nM). Northern-blot analysis showed a significant increase in native receptor mRNA expression in 3′-UTR decoy-expressing cells, confirming the role of 3′-UTR in mRNA destabilization. Compared with vehicle control, AngII induced DNA and protein synthesis in wild-type WB as measured by [3H]thymidine and [3H]leucine incorporation respectively. Activation of [3H]thymidine and [3H]leucine correlated with a significant increase in cell number (cellular hyperplasia). In these cells, AngII stimulated GTPase activity by AT1 receptor coupling with G-protein αi. We also delineated that functional coupling of AT1A receptor with G-protein αi is an essential mechanism for AngII-mediated cellular hyperplasia in WB by specifically blocking G-protein αi activation. In contrast with wild-type cells, stable expression of the 3′-UTR ‘decoy’ produced AngII-stimulated protein synthesis and cellular hypertrophy as demonstrated by a significant increase in [3H]leucine incorporation and no increase in [3H]thymidine incorporation and cell number. Furthermore, [125I]AngII cross-linking and immunoprecipitation studies using specific G-protein α antibodies showed that in wild-type cells, the AT1A receptor coupled with G-protein αi, whereas in cells expressing the 3′-UTR ‘decoy’, the AT1A receptor coupled with G-protein αq. These findings indicate that the 3′-UTR-mediated changes in receptor function may be mediated in part by a switch from G-protein αi to G-protein αq coupling of the receptor. Our results suggest that the 3′-UTR-mediated post-transcriptional modification of the AT1A receptor is critical for regulating tissue-specific receptor functions.

Histamine and inositol phosphate accumulation in endothelium: cAMP and a G protein

1989 ◽  
Vol 257 (4) ◽  
pp. L259-L264 ◽  
Author(s):  
M. R. Carson ◽  
S. S. Shasby ◽  
D. M. Shasby
Keyword(s):  
Endothelial Cells ◽  
Intracellular Calcium ◽  
G Protein ◽  
Inositol Phosphate ◽  
Umbilical Vein ◽  
Microvascular Permeability ◽  
Alpha Toxin ◽  
H1 Receptor ◽  
Phosphate Accumulation ◽  
Inositol Phosphate Accumulation

Histamine increases microvascular permeability through a calcium-dependent process, and histamine occupancy of the H1-receptor increases calcium in cultured endothelial cells. Agents that increase adenosine 3',5'-cyclic monophosphate (cAMP) in endothelial cells prevent the in vivo increase in microvascular permeability that follows histamine exposure. In the current experiments, histamine occupancy of the H1-receptor increased the flux of albumin across monolayers of cultured human umbilical vein endothelial cells (HUVEC). This was prevented by pretreating the cells with theophylline, forskolin, and 8-bromo-cAMP (BrcAMP), which also decreased the flux of albumin across control monolayers. Exposing the cells to histamine increased inositol phosphate accumulation in the cells, and this was prevented by the H1-antagonist pyrilamine but not by theophylline, forskolin, and BrcAMP. Exposing the cells to histamine increased intracellular calcium measured with fura-2. The increase in cell calcium was prevented by pyrilamine but not by pretreatment with theophylline, forskolin, and BrcAMP. When endogenous cell GTP was depleted by permeabilizing the membranes of the endothelial cells with Staphylococcus aureus alpha-toxin, histamine-stimulated inositol phosphate accumulation was enhanced with addition of GTP but not with addition of GDP to the buffer. Addition of GTP alone to the buffer did not increase inositol phosphate accumulation in alpha-toxin-treated cells. Histamine stimulates inositol phosphate accumulation in HUVEC via a G protein. Inhibition of the edemagenic effects of histamine by cAMP does not occur by interrupting this signal transduction pathway between the binding of histamine to its receptor and the increase in intracellular calcium.

scholarly journals Analysis of Missense Variants in the Human Histamine Receptor Family Reveals Increased Constitutive Activity of E4106.30×30K Variant in the Histamine H1 Receptor

2021 ◽  
Vol 22 (7) ◽  
pp. 3702
Author(s):  
Xiaoyuan Ma ◽  
Marta Arimont Segura ◽  
Barbara Zarzycka ◽  
Henry F. Vischer ◽  
Rob Leurs
Keyword(s):  
G Protein ◽  
Missense Variant ◽  
Histamine Receptor ◽  
Structural Basis ◽  
Constitutive Activity ◽  
Wild Type ◽  
H1 Receptor ◽  
Histamine H1 Receptor ◽  
Missense Variants ◽  
Receptor Family

The Exome Aggregation Consortium has collected the protein-encoding DNA sequences of almost 61,000 unrelated humans. Analysis of this dataset for G protein-coupled receptor (GPCR) proteins (available at GPCRdb) revealed a total of 463 naturally occurring genetic missense variations in the histamine receptor family. In this research, we have analyzed the distribution of these missense variations in the four histamine receptor subtypes concerning structural segments and sites important for GPCR function. Four missense variants R1273.52×52H, R13934.57×57H, R4096.29×29H, and E4106.30×30 K, were selected for the histamine H1 receptor (H1R) that were hypothesized to affect receptor activity by interfering with the interaction pattern of the highly conserved D(E)RY motif, the so-called ionic lock. The E4106.30×30 K missense variant displays higher constitutive activity in G protein signaling as compared to wild-type H1R, whereas the opposite was observed for R1273.52×52H, R13934.57×57H, and R4096.29×29H. The E4106.30×30 K missense variant displays a higher affinity for the endogenous agonist histamine than wild-type H1R, whereas antagonist affinity was not affected. These data support the hypothesis that the E4106.30×30 K mutation shifts the equilibrium towards active conformations. The study of these selected missense variants gives additional insight into the structural basis of H1R activation and, moreover, highlights that missense variants can result in pharmacologically different behavior as compared to wild-type receptors and should consequently be considered in the drug discovery process.

scholarly journals Structure of the human histamine H1 receptor gene

1998 ◽  
Vol 335 (3) ◽  
pp. 663-670 ◽  
Author(s):  
Marianne D. DE BACKER ◽  
Inge LOONEN ◽  
Peter VERHASSELT ◽  
Jean-Marc NEEFS ◽  
Walter H. M.L. LUYTEN
Keyword(s):  
G Protein ◽  
Untranslated Region ◽  
Receptor Gene ◽  
Promoter Sequence ◽  
Start Codon ◽  
Luciferase Reporter ◽  
H1 Receptor ◽  
Histamine H1 Receptor ◽  
G Protein Coupled ◽  
Human Histamine

Histamine H1 receptor expression has been reported to change in disorders such as allergic rhinitis, autoimmune myocarditis, rheumatoid arthritis and atherosclerosis. Here we report the isolation and characterization of genomic clones containing the 5´ flanking (regulatory) region of the human histamine H1 receptor gene. An intron of approx. 5.8 kb was identified in the 5´ untranslated region, which suggests that an entire subfamily of G-protein-coupled receptors may contain an intron immediately upstream of the start codon. The transcription initiation site was mapped by 5´ rapid amplification of cDNA ends to a region 6.2 kb upstream of the start codon. Immediately upstream of the transcription start site a fragment of 1.85 kb was identified that showed promoter activity when placed upstream of a luciferase reporter gene and transiently transfected into cells expressing the histamine H1 receptor. The promoter sequence shares a number of characteristics with the promoter sequences of other G-protein-coupled receptor encoding genes, including binding sites for several transcription factors, and the absence of TATA and CAAT sequences at the appropriate locations. The promoter sequence described here differs from that reported previously [Fukui, Fujimoto, Mizuguchi, Sakamoto, Horio, Takai, Yamada and Ito (1994) Biochem. Biophys. Res. Commun. 201, 894–901] because the reported genomic clone was chimaeric. Furthermore our study provides evidence that the 3´ untranslated region of the H1 receptor mRNA is much longer than previously accepted. Together, these findings provide a complete view of the structure of the human histamine H1 receptor gene. Both the coding region of the H1 receptor gene and its promoter region were independently mapped to chromosome 3p25.

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