@article{AgarwalYangRiceetal.2014, author = {Agarwal, Shailesh R. and Yang, Pei-Chi and Rice, Monica and Singer, Cherie A. and Nikolaev, Viacheslav O. and Lohse, Martin J. and Clancy, Colleen E. and Harvey, Robert D.}, title = {Role of Membrane Microdomains in Compartmentation of cAMP Signaling}, series = {PLOS ONE}, volume = {9}, journal = {PLOS ONE}, number = {4}, issn = {1932-6203}, doi = {10.1371/journal.pone.0095835}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-116673}, pages = {e95835}, year = {2014}, abstract = {Spatially restricting cAMP production to discrete subcellular locations permits selective regulation of specific functional responses. But exactly where and how cAMP signaling is confined is not fully understood. Different receptors and adenylyl cyclase isoforms responsible for cAMP production are not uniformly distributed between lipid raft and non-lipid raft domains of the plasma membrane. We sought to determine the role that these membrane domains play in organizing cAMP responses in HEK293 cells. The freely diffusible FRET-based biosensor Epac2-camps was used to measure global cAMP responses, while versions of the probe targeted to lipid raft (Epac2-MyrPalm) and non-raft (Epac2-CAAX) domains were used to monitor local cAMP production near the plasma membrane. Disruption of lipid rafts by cholesterol depletion selectively altered cAMP responses produced by raft-associated receptors. The results indicate that receptors associated with lipid raft as well as non-lipid raft domains can contribute to global cAMP responses. In addition, basal cAMP activity was found to be significantly higher in non-raft domains. This was supported by the fact that pharmacologic inhibition of adenylyl cyclase activity reduced basal cAMP activity detected by Epac2-CAAX but not Epac2-MyrPalm or Epac2-camps. Responses detected by Epac2-CAAX were also more sensitive to direct stimulation of adenylyl cyclase activity, but less sensitive to inhibition of phosphodiesterase activity. Quantitative modeling was used to demonstrate that differences in adenylyl cyclase and phosphodiesterase activities are necessary but not sufficient to explain compartmentation of cAMP associated with different microdomains of the plasma membrane.}, language = {en} } @article{LohseKlotzSchwabe1991, author = {Lohse, Martin J. and Klotz, Karl-Norbert and Schwabe, Ulrich}, title = {Mechanism of A2 adenosine receptor activation. I. Blockade of A2 adenosine receptors by photoaffinity labeling}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-86073}, year = {1991}, abstract = {It has previously been shown that covalent incorporation of the photoreactive adenosine derivative (R)-2-azido-N6-p-hydroxyphenytisopropyladenosine [(R)-AHPIA] into the A, adenosine receptor of intact fat cells leads to a persistent activation of this receptor, resulting in a reduction of celular cAMP Ieveis [Mol. Pharmacol. 30:403-409 (1986)]. In contrast, covalent incorporation of (R)-AHPIA into human platelet membranes, which contain only stimulatory A2 adenosine receptors, reduces adenytate cyclase Stimulation via these receptors. This effect of (R)-AHPIA is specific for the A2 receptor and can be prevented by the adenosine receptor antagonist theophylline. Binding studies in-dicate that up to 90\% of A2 receptors can be blocked by photoincorporation of (R)-AHPIA. However, the remaining 10-20\% of A2 receptors are sufficient to mediate an adenylate cyclase Stimulation of up to SOOk of the control value. Similarly, the activation via these 10-20\% of receptors occurs with a halflife that is only 2 times Ionger than that in control membranes. This indicates the presence of a receptor reserve, with respect to both the extent and the rate of adenytate cyclase Stimulation. These observations require a modification of the models of receptor-adenytate cyclase coupling, which is described in the accompanying paper [Mol. Pharmacol. 39:524-530 (1991)].}, subject = {Adenosinrezeptor}, language = {en} } @incollection{LohseKlotzSchwabe1985, author = {Lohse, Martin J. and Klotz, Karl-Norbert and Schwabe, Ulrich}, title = {Effects of barbiturates on A1 adenosine receptors of rat brain}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-70100}, publisher = {Universit{\"a}t W{\"u}rzburg}, year = {1985}, abstract = {Barbiturates inhibit binding of radioligands to A 1(Ri) adenosine receptors of rat brain membranes. This inhibition is dose-dependent and stereospecific and occurs in the range of pharmacologically active concentrations. The displacement of radiolabelled A1antagonists by barbiturates is not modified by GTP, indicating that barbiturates might act as antagonists at this receptor. This action of barbiturates does not seem to be related to the binding of barbiturates to plasma membranes, as the latter process has different characteristics. Barbiturates also inhibit the binding of radioligands to solubilized A1receptors, and saturation and kinetic experiments suggest that this is due to a competitive antagonism. These results indicate that barbiturates interact with the recognition site of the A1adenosine receptor.}, subject = {Barbiturat}, language = {en} } @incollection{LohseKlotzSchwabe1987, author = {Lohse, Martin J. and Klotz, Karl-Norbert and Schwabe, Ulrich}, title = {Functional characterization of A1 adenoosine receptors by photoaffinity labelling}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-86097}, publisher = {Universit{\"a}t W{\"u}rzburg}, year = {1987}, abstract = {The ligand-binding subunit ofthe A1 adenosine receptor has been identified in membranes with the photoaffinity Iabel R-2-azido-N6-p-hydroxyphenylisopropyladenosine (R-AHPIA). Covalent labelling ofthe A1 receptor can also be achieved in intact cells. The dissociation of the radioiodinated label (1251-AHPIA) from isolated rat fat cells was incomplete after UV irradiation, leaving about 20°/o of irreversible specific binding. Such covalent labelling of the receptor led to a concentration-dependent reduction of cellular cyclic AMP levels. This persistent effect of covalent labeHing occurred with an IC50 value of 9 nM, as compared to an IC50 value of 0.9 nM for the direct reduction of cyclic AMP Ievels by the ligand. The difference in the IC5o values can be explained by assuming spare receptors. This hypothesis was verified in binding studies using [ 3HJPIA as a radioligand. R-AHPIA inhibited binding of [3H)PIA to intact fat cells with a K1 value of about 20 nM, which is about 20 tim es high er than the corresponding IC50 value of cyclic AMP reduction. These data show that the A1 receptor is activated according to the occupancy theory. The high sensitivity of the activation in intact ceJis is due to a large number of spare receptors.}, subject = {Adenosinrezeptor}, language = {en} } @incollection{LohseKlotzMaureretal.1990, author = {Lohse, Martin J. and Klotz, Karl-Norbert and Maurer, K. and Ott, I. and Schwabe, Ulrich}, title = {Effects of adenosine on mast cells}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-86101}, publisher = {Universit{\"a}t W{\"u}rzburg}, year = {1990}, abstract = {No abstract available}, subject = {Adenosin}, language = {en} } @article{LohseKlotzSchwabe1986, author = {Lohse, Martin J. and Klotz, Karl-Norbert and Schwabe, Ulrich}, title = {Effectes of temperature and membrane phase transitions on ligand binding to a2-receptors of human platelets}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-86023}, year = {1986}, abstract = {The binding of agonists and antagonists to a2-adrenergic receptors of human platelets was studied. The receptors showed homogeneaus affinities for antagonists but two affinity states for the agonist (-)-epinephrine, which were modulated by guanine nucleotides. Van't Hoffplots of antagonist binding had a break point at about 18° and considerable diversity between 18° and 0°. Agonist binding to both affinity states showed a similar break point; agonist binding to the high affinity state was characterized by a large entropy component compared to the low affinity state. This entropy component was reduced at higher concentrations of sodium, indicating that it may be due to Iiberation of sodium ions. Measurements of the fluorescence of 1-anilin-8-naphthalenesulfonate showed thermotropic phase transitions of theplatelet membranes at about 17°. The transition temperature was decreased to about 12° by addition of 1 0 mM octanoic acid. Octanoic acidalso shifted the break points of the van't Hoffplot of antagonist and low affinity agonist binding from 18° to 12°. High affinity agonist binding, however, remained unchanged. It is concluded that agonist-specific thennodynamic characteristics of ligand binding to a2-receptors of human platelets can only be investigated by regarding differences between high and low affinity agonist binding. These differences include an entropy increase upon Iigand binding, which is in part due to enhanced liberation of sodium ions, and a loss of sensitivity to fluidity changes in the outer layer of the plasma membrane.}, subject = {Molekularpharmakologie}, language = {en} } @article{LohseKlotzSchwabe1986, author = {Lohse, Martin J. and Klotz, Karl-Norbert and Schwabe, Ulrich}, title = {Agonist photoaffinity labeling of A1 adenosine receptors: Persistent activation reveals spare receptors}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-87966}, year = {1986}, abstract = {No abstract available.}, subject = {Pharmazie}, language = {en} } @article{SchoeneggeGallionPicardetal.2017, author = {Sch{\"o}negge, Anne-Marie and Gallion, Jonathan and Picard, Louis-Philippe and Wilkins, Angela D. and Le Gouill, Christian and Audet, Martin and Stallaert, Wayne and Lohse, Martin J. and Kimmel, Marek and Lichtarge, Olivier and Bouvier, Michel}, title = {Evolutionary action and structural basis of the allosteric switch controlling β\(_2\)AR functional selectivity}, series = {Nature Communications}, volume = {8}, journal = {Nature Communications}, doi = {10.1038/s41467-017-02257-x}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-172268}, year = {2017}, abstract = {Functional selectivity of G-protein-coupled receptors is believed to originate from ligand-specific conformations that activate only subsets of signaling effectors. In this study, to identify molecular motifs playing important roles in transducing ligand binding into distinct signaling responses, we combined in silico evolutionary lineage analysis and structure-guided site-directed mutagenesis with large-scale functional signaling characterization and non-negative matrix factorization clustering of signaling profiles. Clustering based on the signaling profiles of 28 variants of the β\(_2\)-adrenergic receptor reveals three clearly distinct phenotypical clusters, showing selective impairments of either the Gi or βarrestin/endocytosis pathways with no effect on Gs activation. Robustness of the results is confirmed using simulation-based error propagation. The structural changes resulting from functionally biasing mutations centered around the DRY, NPxxY, and PIF motifs, selectively linking these micro-switches to unique signaling profiles. Our data identify different receptor regions that are important for the stabilization of distinct conformations underlying functional selectivity.}, language = {en} } @article{SchihadaVandenabeeleZabeletal.2018, author = {Schihada, Hannes and Vandenabeele, Sylvie and Zabel, Ulrike and Frank, Monika and Lohse, Martin J. and Maiellaro, Isabella}, title = {A universal bioluminescence resonance energy transfer sensor design enables high-sensitivity screening of GPCR activation dynamics}, series = {Communications Biology}, volume = {1}, journal = {Communications Biology}, number = {105}, doi = {10.1038/s42003-018-0072-0}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-228592}, pages = {1-8}, year = {2018}, abstract = {G-protein-coupled receptors (GPCRs) represent one of the most important classes of drug targets. The discovery of new GCPR therapeutics would greatly benefit from the development of a generalizable high-throughput assay to directly monitor their activation or de-activation. Here we screened a variety of labels inserted into the third intracellular loop and the C-terminus of the alpha(2 Lambda)-adrenergic receptor and used fluorescence (FRET) and bioluminescence resonance energy transfer (BRET) to monitor ligand-binding and activation dynamics. We then developed a universal intramolecular BRET receptor sensor design to quantify efficacy and potency of GPCR ligands in intact cells and real time. We demonstrate the transferability of the sensor design by cloning beta(2)-adrenergic and PTH1-receptor BRET sensors and monitored their efficacy and potency. For all biosensors, the Z factors were well above 0.5 showing the suitability of such design for microtiter plate assays. This technology will aid the identification of novel types of GPCR ligands.}, language = {en} } @article{PaisdziorDimitriouSchoepeetal.2020, author = {Paisdzior, Sarah and Dimitriou, Ioanna Maria and Sch{\"o}pe, Paul Curtis and Annibale, Paolo and Scheerer, Patrick and Krude, Heiko and Lohse, Martin J. and Biebermann, Heike and K{\"u}hnen, Peter}, title = {Differential signaling profiles of MC4R mutations with three different ligands}, series = {International Journal of Molecular Sciences}, volume = {21}, journal = {International Journal of Molecular Sciences}, number = {4}, issn = {1422-0067}, doi = {10.3390/ijms21041224}, url = {http://nbn-resolving.de/urn:nbn:de:bvb:20-opus-285108}, year = {2020}, abstract = {The melanocortin 4 receptor (MC4R) is a key player in hypothalamic weight regulation and energy expenditure as part of the leptin-melanocortin pathway. Mutations in this G protein coupled receptor (GPCR) are the most common cause for monogenetic obesity, which appears to be mediated by changes in the anorectic action of MC4R via G\(_S\)-dependent cyclic adenosine-monophosphate (cAMP) signaling as well as other signaling pathways. To study potential bias in the effects of MC4R mutations between the different signaling pathways, we investigated three major MC4R mutations: a G\(_S\) loss-of-function (S127L) and a G\(_S\) gain-of-function mutant (H158R), as well as the most common European single nucleotide polymorphism (V103I). We tested signaling of all four major G protein families plus extracellular regulated kinase (ERK) phosphorylation and β-arrestin2 recruitment, using the two endogenous agonists, α- and β-melanocyte stimulating hormone (MSH), along with a synthetic peptide agonist (NDP-α-MSH). The S127L mutation led to a full loss-of-function in all investigated pathways, whereas V103I and H158R were clearly biased towards the G\(_{q/11}\) pathway when challenged with the endogenous ligands. These results show that MC4R mutations can cause vastly different changes in the various MC4R signaling pathways and highlight the importance of a comprehensive characterization of receptor mutations.}, language = {en} }