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Gerritjan Koomen - One of the best experts on this subject based on the ideXlab platform.
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n6 cyclopentyl 2 3 phenylaminocarbonyltriazene 1 yl adenosine tcpa a very selective agonist with high affinity for the human adenosine a1 receptor
Journal of Medicinal Chemistry, 2003Co-Authors: Margot W Beukers, Martin J Wanner, Jacobien K Von Frijtag Drabbe Kunzel, Elisabeth Klaasse, And A P Ijzerman, Gerritjan KoomenAbstract:Four subtypes of adenosine receptors are currently known, that is, A1, A2A, A2B, and A3 receptors. Interestingly, quite substantial species differences exist especially between human and rat A3 receptors. As a result, ligands such as CCPA, which are very selective for the rat A1 receptor versus the human A3 receptor, are substantially less selective when the human A1 and A3 receptors are compared. New 2-substituted and 2,N6-disubstituted Adenosines were synthesized, and their affinities for the human adenosine A1, A2A, A2B, and A3 receptors were determined. Although large substituents on the C2-position are generally thought to yield adenosine A2A receptor selective ligands, the reported series of 2-triazeno-substituted Adenosines had a very high affinity for the A1 receptor. For example, 2-(3-phenylaminocarbonyltriazene-1-yl)adenosine had an affinity of 6.1 ± 1.3 nM for the human adenosine A1 receptor. Introduction of a diphenethyl substituent at the N6-position of this compound resulted in a high-affini...
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n6 cyclopentyl 2 3 phenylaminocarbonyltriazene 1 yl adenosine tcpa a very selective agonist with high affinity for the human adenosine a1 receptor
Journal of Medicinal Chemistry, 2003Co-Authors: Margot W Beukers, Martin J Wanner, Elisabeth Klaasse, And A P Ijzerman, Jacobien K Von Frijtag Drabbe Kunzel, Gerritjan KoomenAbstract:Four subtypes of adenosine receptors are currently known, that is, A(1), A(2A), A(2B), and A(3) receptors. Interestingly, quite substantial species differences exist especially between human and rat A(3) receptors. As a result, ligands such as CCPA, which are very selective for the rat A(1) receptor versus the human A(3) receptor, are substantially less selective when the human A(1) and A(3) receptors are compared. New 2-substituted and 2,N(6)-disubstituted Adenosines were synthesized, and their affinities for the human adenosine A(1), A(2A), A(2B), and A(3) receptors were determined. Although large substituents on the C2-position are generally thought to yield adenosine A(2A) receptor selective ligands, the reported series of 2-triazeno-substituted Adenosines had a very high affinity for the A(1) receptor. For example, 2-(3-phenylaminocarbonyltriazene-1-yl)adenosine had an affinity of 6.1 +/- 1.3 nM for the human adenosine A(1) receptor. Introduction of a diphenethyl substituent at the N(6)-position of this compound resulted in a high-affinity agonist, 3.1 +/- 0.9 nM, for the human adenosine A(1) receptor with 316- and 45-fold selectivity versus the human A(2A) and human A(3) receptors, respectively. The most selective, high-affinity human adenosine A(1) receptor agonist was the disubstituted compound N(6)-cyclopentyl-2-(3-phenylaminocarbonyltriazene-1-yl)adenosine (TCPA). TCPA had an affinity of 2.8 +/- 0.8 nM for the human adenosine A(1) receptor and was 75-fold and 214-fold selective versus the human A(2A) and human A(3) receptors, respectively. In addition, TCPA was a full agonist and inhibited the forskolin-induced cAMP production of CHO cells stably transfected with the human adenosine A(1) receptor with an IC(50) of 1.5 +/- 0.5 nM.
Kenneth A Jacobson - One of the best experts on this subject based on the ideXlab platform.
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convergent synthesis of 2 thioether substituted n methanocarba Adenosines as purine receptor agonists
RSC Advances, 2021Co-Authors: Rama R Suresh, Russell B Poe, Baorui Lin, Ryan Campbell, Zhanguo Gao, Theodore E Liston, Kiran S Toti, Kenneth A JacobsonAbstract:A linear route has been used to prepare (N)-methanocarba-nucleoside derivatives, which serve as purine receptor ligands having a pre-established, receptor-preferred conformation. To introduce this rigid ribose substitute, a Mitsunobu reaction of a [3.1.0]bicyclohexane 5′-trityl intermediate 3 with a nucleobase is typically followed by functional group modifications. We herein report an efficient scalable convergent synthesis for 2-substituted (N)-methanocarba-Adenosines, which were demonstrated to bind to the A3 adenosine receptor. The adenine moiety was pre-functionalized with 2-thioethers and other groups before coupling to the bicyclic precursor (3) as a key step to facilitate a high yield Mitsunobu product. This new approach provided the (N)-methanocarba-Adenosines in moderate to good yield, which effectively increased the overall yield compared to a linear synthesis and conserved a key intermediate 3 (a product of nine sequential steps). The generality of this convergent synthesis, which is suitable as an optimized preclinical synthetic route, was demonstrated with various 2-thioether and 2-methoxy substituents.
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structural sweet spot for a1 adenosine receptor activation by truncated n methanocarba nucleosides receptor docking and potent anticonvulsant activity
Journal of Medicinal Chemistry, 2012Co-Authors: Dilip K Tosh, Zhanguo Gao, Silvia Paoletta, Francesca Deflorian, Khai Phan, Steven M Moss, Xiaohui Jiang, Kenneth A JacobsonAbstract:A1 adenosine receptor (AR) agonists display antiischemic and antiepileptic neuroprotective activity, but peripheral cardiovascular side effects impeded their development. SAR study of N6-cycloalkylmethyl 4′-truncated (N)-methanocarba-Adenosines identified 10 (MRS5474, N6-dicyclopropylmethyl, Ki = 47.9 nM) as a moderately A1AR-selective full agonist. Two stereochemically defined N6-methynyl group substituents displayed narrow SAR; groups larger than cyclobutyl greatly reduced AR affinity, and those larger or smaller than cyclopropyl reduced A1AR selectivity. Nucleoside docking to A1AR homology model characterized distinct hydrophobic cyclopropyl subpockets, the larger “A” forming contacts with Thr270 (7.35), Tyr271 (7.36), Ile274 (7.39), and carbon chains of glutamates (EL2) and the smaller subpocket “B” forming contacts between TM6 and TM7. 10 suppressed minimal clonic seizures (6 Hz mouse model) without typical rotarod impairment of A1AR agonists. Truncated nucleosides, an appealing preclinical approach,...
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Structural Sweet Spot for A1 Adenosine Receptor Activation by Truncated (N)-Methanocarba Nucleosides: Receptor Docking and Potent Anticonvulsant Activity
2012Co-Authors: Dilip K. Tosh, Zhanguo Gao, Silvia Paoletta, Francesca Deflorian, Khai Phan, Xiaohui Jiang, Steven M. Moss, Kenneth A JacobsonAbstract:A1 adenosine receptor (AR) agonists display antiischemic and antiepileptic neuroprotective activity, but peripheral cardiovascular side effects impeded their development. SAR study of N6-cycloalkylmethyl 4′-truncated (N)-methanocarba-Adenosines identified 10 (MRS5474, N6-dicyclopropylmethyl, Ki = 47.9 nM) as a moderately A1AR-selective full agonist. Two stereochemically defined N6-methynyl group substituents displayed narrow SAR; groups larger than cyclobutyl greatly reduced AR affinity, and those larger or smaller than cyclopropyl reduced A1AR selectivity. Nucleoside docking to A1AR homology model characterized distinct hydrophobic cyclopropyl subpockets, the larger “A” forming contacts with Thr270 (7.35), Tyr271 (7.36), Ile274 (7.39), and carbon chains of glutamates (EL2) and the smaller subpocket “B” forming contacts between TM6 and TM7. 10 suppressed minimal clonic seizures (6 Hz mouse model) without typical rotarod impairment of A1AR agonists. Truncated nucleosides, an appealing preclinical approach, have more druglike physicochemical properties than other A1AR agonists. Thus, we identified highly restricted regions for substitution around N6 suitable for an A1AR agonist with anticonvulsant activity
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n6 substituted adenosine derivatives selectivity efficacy and species differences at a3 adenosine receptors
Biochemical Pharmacology, 2003Co-Authors: Zhanguo Gao, Joshua B Blaustein, Ariel S Gross, Neli Melman, Kenneth A JacobsonAbstract:The activation of the human A3 adenosine receptor (AR) by a wide range of N6-substituted adenosine derivatives was studied in intact CHO cells stably expressing this receptor. Selectivity of binding at rat and human ARs was also determined. Among N6-alkyl substitutions, small N6-alkyl groups were associated with selectivity for human A3ARs vs. rat A3ARs, and multiple points of branching were associated with decreased hA3AR efficacy. N6-Cycloalkyl-substituted Adenosines were full (≤5 carbons) or partial (≥6 carbons) hA3AR agonists. N6-(endo-Norbornyl)adenosine 13 was the most selective for both rat and human A1ARs. Numerous N6-arylmethyl analogues, including substituted benzyl, tended to be more potent in binding to A1 and A3 vs. A2AARs (with variable degrees of partial to full A3AR agonisms). A chloro substituent decreased the efficacy depending on its position on the benzyl ring. The A3AR affinity and efficacy of N6-arylethyl Adenosines depended highly on stereochemistry, steric bulk, and ring constraints. Stereoselectivity of binding was demonstrated for N6-(R-1-phenylethyl)adenosine vs. N6-(S-1-phenylethyl)adenosine, as well as for the N6-(1-phenyl-2-pentyl)adenosine, at the rat, but not human A3AR. Interestingly, DPMA, a potent agonist for the A2AAR (Ki = 4 nM), was demonstrated to be a moderately potent antagonist for the human A3AR (Ki = 106 nM). N6-[(1S,2R)-2-Phenyl-1-cyclopropyl]adenosine 48 was 1100-fold more potent in binding to human (Ki = 0.63 nM) than rat A3ARs. Dual acting A1/A3 agonists (N6-3-chlorobenzyl- 29, N6-(S-1-phenylethyl)- 39, and 2-chloro-N6-(R-phenylisopropyl)adenosine 53) might be useful for cardioprotection.
Margot W Beukers - One of the best experts on this subject based on the ideXlab platform.
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n6 cyclopentyl 2 3 phenylaminocarbonyltriazene 1 yl adenosine tcpa a very selective agonist with high affinity for the human adenosine a1 receptor
Journal of Medicinal Chemistry, 2003Co-Authors: Margot W Beukers, Martin J Wanner, Jacobien K Von Frijtag Drabbe Kunzel, Elisabeth Klaasse, And A P Ijzerman, Gerritjan KoomenAbstract:Four subtypes of adenosine receptors are currently known, that is, A1, A2A, A2B, and A3 receptors. Interestingly, quite substantial species differences exist especially between human and rat A3 receptors. As a result, ligands such as CCPA, which are very selective for the rat A1 receptor versus the human A3 receptor, are substantially less selective when the human A1 and A3 receptors are compared. New 2-substituted and 2,N6-disubstituted Adenosines were synthesized, and their affinities for the human adenosine A1, A2A, A2B, and A3 receptors were determined. Although large substituents on the C2-position are generally thought to yield adenosine A2A receptor selective ligands, the reported series of 2-triazeno-substituted Adenosines had a very high affinity for the A1 receptor. For example, 2-(3-phenylaminocarbonyltriazene-1-yl)adenosine had an affinity of 6.1 ± 1.3 nM for the human adenosine A1 receptor. Introduction of a diphenethyl substituent at the N6-position of this compound resulted in a high-affini...
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n6 cyclopentyl 2 3 phenylaminocarbonyltriazene 1 yl adenosine tcpa a very selective agonist with high affinity for the human adenosine a1 receptor
Journal of Medicinal Chemistry, 2003Co-Authors: Margot W Beukers, Martin J Wanner, Elisabeth Klaasse, And A P Ijzerman, Jacobien K Von Frijtag Drabbe Kunzel, Gerritjan KoomenAbstract:Four subtypes of adenosine receptors are currently known, that is, A(1), A(2A), A(2B), and A(3) receptors. Interestingly, quite substantial species differences exist especially between human and rat A(3) receptors. As a result, ligands such as CCPA, which are very selective for the rat A(1) receptor versus the human A(3) receptor, are substantially less selective when the human A(1) and A(3) receptors are compared. New 2-substituted and 2,N(6)-disubstituted Adenosines were synthesized, and their affinities for the human adenosine A(1), A(2A), A(2B), and A(3) receptors were determined. Although large substituents on the C2-position are generally thought to yield adenosine A(2A) receptor selective ligands, the reported series of 2-triazeno-substituted Adenosines had a very high affinity for the A(1) receptor. For example, 2-(3-phenylaminocarbonyltriazene-1-yl)adenosine had an affinity of 6.1 +/- 1.3 nM for the human adenosine A(1) receptor. Introduction of a diphenethyl substituent at the N(6)-position of this compound resulted in a high-affinity agonist, 3.1 +/- 0.9 nM, for the human adenosine A(1) receptor with 316- and 45-fold selectivity versus the human A(2A) and human A(3) receptors, respectively. The most selective, high-affinity human adenosine A(1) receptor agonist was the disubstituted compound N(6)-cyclopentyl-2-(3-phenylaminocarbonyltriazene-1-yl)adenosine (TCPA). TCPA had an affinity of 2.8 +/- 0.8 nM for the human adenosine A(1) receptor and was 75-fold and 214-fold selective versus the human A(2A) and human A(3) receptors, respectively. In addition, TCPA was a full agonist and inhibited the forskolin-induced cAMP production of CHO cells stably transfected with the human adenosine A(1) receptor with an IC(50) of 1.5 +/- 0.5 nM.
Peter J Scammells - One of the best experts on this subject based on the ideXlab platform.
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synthesis and biological evaluation of Adenosines with heterobicyclic and polycyclic n6 substituents as adenosine a1 receptor agonists
ChemMedChem, 2012Co-Authors: Joshua Immanuel Gosling, Stephen P Baker, John M Haynes, Michael Kassiou, Colin W Pouton, Lyndon Warfe, Paul J White, Peter J ScammellsAbstract:A concise synthesis of a series of N(6)-substituted Adenosines with bicyclo[3.2.1]octan-6-yl and polycyclic N(6)-substituents has been developed. The adenosine A(1) receptor (A(1)R) affinity and potency of these compounds was initially assessed using competitive binding assays and cyclic adenosine monophosphate (cAMP) accumulation assays in DDT(1) MF-2 cells. The potency and receptor subtype selectivity of selected examples was further evaluated by measuring their effects on cAMP accumulation at all human adenosine receptor subtypes expressed in CHO cells. The results of these assays indicated that all of the synthesised N(6)-substituted Adenosines are full agonists at A(1) R and activate this receptor selectively over the other adenosine receptor subtypes. The two standout compounds in terms of potency were N(6)-(3-thiabicyclo[3.2.1]octan-6-yl)adenosine and N(6)-(cubanylmethyl)adenosine with EC(50) values at human A(1)R of 2.3 nM and 1.1 nM, respectively. The cubanylmethyl derivative in particular proved to be highly receptor subtype selective. These two compounds were further evaluated in a simulated ischaemia model in cultured cardiomyoblasts, where they were found to impart protective effects under hypoxic conditions that resulted in a significant reduction in cell death.
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microwave assisted direct amination rapid access to multi functionalized n6 substituted Adenosines
ChemInform, 2008Co-Authors: Trent D Ashton, Peter J ScammellsAbstract:Analogues of adenosine have a range of interesting biological activities and potential therapeutic applications. A method for the efficient preparation of highly functionalized N6-substituted Adenosines has been developed from the corresponding tert-butyldimethylsilyl-protected inosine. The key step in this procedure is a microwave-assisted amination reaction between an appropriately substituted inosine and an amine in the presence of PyBroP. High yields of desired N6-substituted Adenosines were achieved with hindered amines and the reaction was also found to accommodate a range of substituents on the inosine precursor.
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structure activity relationships of Adenosines with heterocyclic n6 substituents
ChemInform, 2008Co-Authors: Trent D Ashton, Stephen P Baker, Kylee M Aumann, Carl H Schiesser, Peter J ScammellsAbstract:Two series of N(6)-substituted Adenosines with monocyclic and bicyclic N(6) substituents containing a heteroatom were synthesized in good yields. These derivatives were assessed for their affinity ([(3)H]CPX), potency, and intrinsic activity (cAMP accumulation) at the A(1) adenosine receptor in DDT(1) MF-2 cells. In the monocyclic series, the N(6)-tetrahydrofuran-3-yl and thiolan-3-yl Adenosines (1 and 26, respectively) were found to possess similar activities, whereas the corresponding selenium analogue 27 was found to be more potent. A series of nitrogen containing analogues showed varying properties, N(6)-((3R)-1-benzyloxycarbonylpyrrolidin-3-yl)adenosine (30) was the most potent at the A(1)AR; IC(50)=3.2 nM. In the bicyclic series, the effect of a 7-azabicyclo[2.2.1]heptan-2-yl substituent in the N(6)-position was explored. N(6)-(7-Azabicyclo[2.2.1]heptan-2-yl)adenosine (38) proved to be a reasonably potent A(1) agonist (K(i)=51 nM, IC(50)=35 nM) while further substitution on the 7''-nitrogen with tert-butoxycarbonyl (31, IC(50)=2.5 nM) and 2-bromobenzyloxycarbonyl (34, IC(50)=9.0 nM) gave highly potent A(1)AR agonists.
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n6 substituted c5 modified Adenosines as a1 adenosine receptor agonists
Bioorganic & Medicinal Chemistry, 2008Co-Authors: Trent D Ashton, Stephen P Baker, Sally A Hutchinson, Peter J ScammellsAbstract:Abstract Adenosines bearing 5′-modification in conjunction with an N6-substituent have previously been shown to act as partial agonists at the A1 adenosine receptor. Our current work investigates the effect of modifying the 5′-position in conjunction with efficacious bicyclic and tricyclic N6-substituents. Several highly potent agonists for the A1 adenosine receptor were identified; however, all of these compounds behaved as full agonists. In keeping with previous reports, 5′-halogen and 5′-sulfide derivatives of N6-(endo-norborn-2-yl)adenosine were, in general, low nanomolar agonists of the A1 adenosine receptor. The known partial agonist, N6-cyclopentyl-5′-deoxy-5′-ethylthioadenosine (2), also behaved as a full agonist in our assay.
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new 2 n6 disubstituted Adenosines potent and selective a1 adenosine receptor agonists
ChemInform, 2002Co-Authors: Sally A Hutchinson, Stephen P Baker, Peter J ScammellsAbstract:A number of adenosine analogues substituted in the 2- and N6-positions were synthesized and evaluated for affinity, functional potency and intrinsic activity at the A1 and A2A adenosine receptors (AR). Three classes of N6-substituents were tested; norbornen-2-yl (series 1), norborn-2-yl (series 2) and 5,6-epoxynorborn-2-yl (series 3). The halogens; fluoro, bromo, and iodo were evaluated as C-2 substituents. All compounds showed relatively high affinity (nanomolar) for the A1AR and high potency for inhibiting (−)isoproterenol-stimulated cAMP accumulation in hamster smooth muscle DDT1 MF-2 cells with the 2-fluoro derivatives from each series having the highest affinity. All of the derivatives showed the same intrinsic activity as CPA. At the A2AAR, all of the derivatives showed relatively low affinity and potency (micromolar) for stimulating cAMP accumulation in rat pheochromocytoma PC-12 cells. The intrinsic activity of the derivatives compared to CGS 21680 was dependent upon the halogen substituent in the C-2 position with most showing partial agonist activity. Of particular interest is 2-iodo-N6-(2S-endo-norborn-2-yl)adenosine (5e), which is over 100-fold selective for the A1AR, is a full agonist at this receptor subtype and has no detectable agonist activity at the A2AAR.
Peter H Seeburg - One of the best experts on this subject based on the ideXlab platform.
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liver disintegration in the mouse embryo caused by deficiency in the rna editing enzyme adar1
Journal of Biological Chemistry, 2004Co-Authors: Jochen C Hartner, Miyoko Higuchi, Carolin Schmittwolf, Andreas Kispert, Albrecht M Muller, Peter H SeeburgAbstract:ADAR1 (adenosine deaminase acting on RNA-1) is widely expressed in mammals, but its biological role is unknown. We show here by gene targeting that ADAR1 selectively edits in vivo two of five closely spaced Adenosines in the serotonin 5-hydroxytryptamine subtype 2C receptor pre-mRNA of nervous tissue; and hence, site-selective adenosine-to-inosine editing is indeed a function of ADAR1. Remarkably, homozygosity for two different null alleles of ADAR1 caused a consistent embryonic phenotype appearing early at embryonic day 11 and leading to death between embryonic days 11.5 and 12.5. This phenotype manifests a rapidly disintegrating liver structure, along with severe defects in definitive hematopoiesis, encompassing both erythroid and myeloid/granuloid progenitors as well as spleen colony-forming activity from the aorta-gonad-mesonephros region and fetal liver. Probably as a consequence of these developmental impairments, ADAR1-deficient embryonic stem cells failed to contribute to liver, bone marrow, spleen, thymus, and blood in adult chimeric mice. Thus, ADAR1 subserves critical steps in developing non-nervous tissue, which are likely to include transcript editing.
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red2 a brain specific member of the rna specific adenosine deaminase family
Journal of Biological Chemistry, 1996Co-Authors: Thorsten Melcher, Stefan Maas, Miyoko Higuchi, Rolf Sprengel, Anne Herb, Peter H SeeburgAbstract:Abstract The mammalian RNA-specific adenosine deaminases DRADA/dsRAD (alias ADAR) and RED1 (alias ADARB1) have been implicated in the site-selective editing of brain-expressed pre-mRNAs for glutamate receptor subunits and of antigenomic RNA of hepatitis delta virus. These enzymes are expressed in many if not all tissues, predicting an as yet unappreciated significance for adenosine deamination-mediated recoding of gene transcripts in the mammalian organism. We now report the molecular cloning of cDNA for RED2 (alias ADARB2), a third member of the RNA-specific adenosine deaminase family in the rodent. RED2 is closely sequence-related to RED1 but appears to be expressed only in the brain, where expression is widespread reaching highest levels in olfactory bulb and thalamus. RED2 further differs from RED1 in having a 54-residue amino-terminal extension which includes an arginine-rich motif. Different from DRADA and RED1, recombinantly expressed RED2 did not deaminate Adenosines in extended synthetic dsRNA or in GluR-B pre-mRNA. However, a chimera of RED1 and RED2 edited the GluR-B Q/R and R/G sites with moderate efficiency. Our data suggest that RED2 may edit brain-specific transcripts with distinct structural features.
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structural requirements for rna editing in glutamate receptor pre mrnas by recombinant double stranded rna adenosine deaminase
Journal of Biological Chemistry, 1996Co-Authors: Stefan Maas, Miyoko Higuchi, Peter H Seeburg, Thorsten Melcher, Walter Keller, Anne Herb, Sabine Krause, Mary A OconnellAbstract:Abstract Pre-mRNAs for brain-expressed ionotropic glutamate receptor subunits undergo RNA editing by site-specific adenosine deamination, which alters codons for molecular determinants of channel function. This nuclear process requires double-stranded RNA structures formed by exonic and intronic sequences in the pre-mRNA and is likely to be catalyzed by an adenosine deaminase that recognizes these structures as a substrate. DRADA, a double-stranded RNA adenosine deaminase, is a candidate enzyme for L-glutamate-activated receptor channel (GluR) pre-mRNA editing. We show here that DRADA indeed edits GluR pre-mRNAs, but that it displays selectivity for certain editing sites. Recombinantly expressed DRADA, both in its full-length form and in an N-terminally truncated version, edited the Q/R site in GluR6 pre-mRNA and the R/G site but not the Q/R site of GluR-B pre-mRNA. This substrate selectivity correlated with the base pairing status and sequence environment of the editing-targeted Adenosines. The Q/R site of GluR-B pre-mRNA was edited by an activity partially purified from HeLa cells and thus differently structured editing sites in GluR pre-mRNAs appear to be substrates for different enzymatic activities.
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editing of α amino 3 hydroxy 5 methylisoxazole 4 propionic acid receptor glur b pre mrna in vitro reveals site selective adenosine to inosine conversion
Journal of Biological Chemistry, 1995Co-Authors: Thorsten Melcher, Stefan Maas, Miyoko Higuchi, Walter Keller, Peter H SeeburgAbstract:Abstract In neurons of the mammalian brain primary transcripts of genes encoding subunits of glutamate receptor channels can undergo RNA editing, leading to altered properties of the transmitter-activated channel. Editing of these transcripts is a nuclear process that targets specific Adenosines and requires a double-stranded RNA structure configured from complementary exonic and intronic sequences. We show here that the two independent editing sites in α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor GluR-B pre-mRNA are edited with positional accuracy by nuclear extract from HeLa cells. Nucleotide analysis by thin layer chromatography of the edited RNA sequences revealed selective adenosine to inosine conversion, most likely reflecting the participation of double-stranded RNA adenosine deaminase. Our results predict the presence of inosine-containing codons in other mammalian mRNAs.
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rna editing of ampa receptor subunit glur b a base paired intron exon structure determines position and efficiency
Cell, 1993Co-Authors: Miyoko Higuchi, Frank Nicolai Single, Martin Kohler, Bernd Sommer, Rolf Sprengel, Peter H SeeburgAbstract:A functionally critical position (Q/R site) of the AMPA receptor subunit GluR-B is controlled by RNA editing that operates in the nucleus, since in brain and clonal cell lines of neural origin, unspliced GluR-B transcripts occur edited in the Q/R site CAG codon and, additionally, in intronic Adenosines. Transfection of GluR-B gene constructs into PC12 cells revealed that the proximal part of the intron downstream of the unedited exonic site is required for Q/R site editing. This intron portion contains an imperfect inverted repeat preceding a 10 nt sequence with exact complementarity to the exon centered on the unedited codon. Single nucleotide substitutions in this short intronic sequence or its exonic complement curtailed Q/R site editing, which was recovered by restoring complementarity in the respective partner strand. Base conversion in the channel-coding region of GluR-B directed by base paired sequences may be executed by a ubiquitous nuclear adenosine deaminase specific for double-stranded RNA.