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Identification of negative residues in the P2X3 ATP receptor ectodomain as structural determinants for desensitization and the Ca2+-sensing modulatory sites.
J Biol Chem 2004; 279(51):53109-15JB

Abstract

On nociceptive neurons, one important mechanism to generate pain signals is the activation of P2X(3) receptors, which are membrane proteins gated by extracellular ATP. In the presence of the agonist, P2X(3) receptors rapidly desensitize and then recover slowly. One unique property of P2X(3) receptors is the recovery acceleration by extracellular Ca(2+) that can play the role of the gain-setter of receptor function only when P2X(3) receptors are desensitized. To study negatively charged sites potentially responsible for this action of Ca(2+), we mutated 15 non-conserved aspartate or glutamate residues in the P2X(3) receptor ectodomain with alanine and expressed such mutated receptors in human embryonic kidney cells studied with patch clamping. Unlike most mutants, D266A (P2X(3) receptor numbering) desensitized very slowly, indicating that this residue is important for generating desensitization. Recovery appeared structurally distinct from desensitization because E111A and D266A had a much faster recovery and D220A and D289A had a much slower one despite their standard desensitization. Furthermore, E161A, E187A, or E270A mutants showed lessened sensitivity to the action of extracellular Ca(2+), suggesting that these determinants were important for the effect of this cation on desensitization recovery. This study is the first report identifying several negative residues in the P2X(3) receptor ectodomain differentially contributing to the general process of receptor desensitization. At least one residue was important to enable the development of rapid desensitization, whereas others controlled recovery from it or the facilitating action of Ca(2+). Thus, these findings outline diverse potential molecular targets to modulate P2X(3) receptor function in relation to its functional state.

Authors+Show Affiliations

Neurobiology Sector, International School for Advanced Studies (SISSA), Via Beirut 4, 34014 Trieste, Italy.

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

15475563

Citation

Fabbretti, Elsa, et al. "Identification of Negative Residues in the P2X3 ATP Receptor Ectodomain as Structural Determinants for Desensitization and the Ca2+-sensing Modulatory Sites." The Journal of Biological Chemistry, vol. 279, no. 51, 2004, pp. 53109-15.
Fabbretti E, Sokolova E, Masten L, et al. Identification of negative residues in the P2X3 ATP receptor ectodomain as structural determinants for desensitization and the Ca2+-sensing modulatory sites. J Biol Chem. 2004;279(51):53109-15.
Fabbretti, E., Sokolova, E., Masten, L., D'Arco, M., Fabbro, A., Nistri, A., & Giniatullin, R. (2004). Identification of negative residues in the P2X3 ATP receptor ectodomain as structural determinants for desensitization and the Ca2+-sensing modulatory sites. The Journal of Biological Chemistry, 279(51), pp. 53109-15.
Fabbretti E, et al. Identification of Negative Residues in the P2X3 ATP Receptor Ectodomain as Structural Determinants for Desensitization and the Ca2+-sensing Modulatory Sites. J Biol Chem. 2004 Dec 17;279(51):53109-15. PubMed PMID: 15475563.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Identification of negative residues in the P2X3 ATP receptor ectodomain as structural determinants for desensitization and the Ca2+-sensing modulatory sites. AU - Fabbretti,Elsa, AU - Sokolova,Elena, AU - Masten,Lara, AU - D'Arco,Marianna, AU - Fabbro,Alessandra, AU - Nistri,Andrea, AU - Giniatullin,Rashid, Y1 - 2004/10/08/ PY - 2004/10/12/pubmed PY - 2005/2/5/medline PY - 2004/10/12/entrez SP - 53109 EP - 15 JF - The Journal of biological chemistry JO - J. Biol. Chem. VL - 279 IS - 51 N2 - On nociceptive neurons, one important mechanism to generate pain signals is the activation of P2X(3) receptors, which are membrane proteins gated by extracellular ATP. In the presence of the agonist, P2X(3) receptors rapidly desensitize and then recover slowly. One unique property of P2X(3) receptors is the recovery acceleration by extracellular Ca(2+) that can play the role of the gain-setter of receptor function only when P2X(3) receptors are desensitized. To study negatively charged sites potentially responsible for this action of Ca(2+), we mutated 15 non-conserved aspartate or glutamate residues in the P2X(3) receptor ectodomain with alanine and expressed such mutated receptors in human embryonic kidney cells studied with patch clamping. Unlike most mutants, D266A (P2X(3) receptor numbering) desensitized very slowly, indicating that this residue is important for generating desensitization. Recovery appeared structurally distinct from desensitization because E111A and D266A had a much faster recovery and D220A and D289A had a much slower one despite their standard desensitization. Furthermore, E161A, E187A, or E270A mutants showed lessened sensitivity to the action of extracellular Ca(2+), suggesting that these determinants were important for the effect of this cation on desensitization recovery. This study is the first report identifying several negative residues in the P2X(3) receptor ectodomain differentially contributing to the general process of receptor desensitization. At least one residue was important to enable the development of rapid desensitization, whereas others controlled recovery from it or the facilitating action of Ca(2+). Thus, these findings outline diverse potential molecular targets to modulate P2X(3) receptor function in relation to its functional state. SN - 0021-9258 UR - https://www.unboundmedicine.com/medline/citation/15475563/Identification_of_negative_residues_in_the_P2X3_ATP_receptor_ectodomain_as_structural_determinants_for_desensitization_and_the_Ca2+_sensing_modulatory_sites_ L2 - http://www.jbc.org/cgi/pmidlookup?view=long&pmid=15475563 DB - PRIME DP - Unbound Medicine ER -