title: Molecular bases of defective signal transduction in the platelet P2Y12 receptor of a patient with congenital bleeding.
abstract: We have identified structural attributes required for signal transduction through a seven-transmembrane-domain receptor. Platelets from a patient (AC) with a congenital bleeding disorder had normal shape change but reduced and reversible aggregation in response to 4 microM ADP, similar to normal platelets with blocked P2Y(12) receptor. The response to 20 microM ADP, albeit still decreased, was more pronounced and was reduced by a P2Y(12) antagonist, indicating some residual receptor function. ADP failed to lower the adenylyl cyclase activity stimulated by prostaglandin E(1) in the patient's platelets, even though the number and affinity of 2-methylthioadenosine 5'-[(33)P]diphosphate-binding sites was normal. Analysis of the patient's P2Y(12) gene revealed a G-to-A transition in one allele, changing the codon for Arg-256 in the sixth transmembrane domain to Gln, and a C-to-T transition in the other allele, changing the codon for Arg-265 in the third extracellular loop to Trp. Neither mutation interfered with receptor surface expression but both altered function, since ADP inhibited the forskolin-induced increase of cAMP markedly less in cells transfected with either mutant P2Y(12) as compared with wild-type receptor. These studies delineate a region of P2Y(12) required for normal function after ADP binding.
authors.affiliation: A. Bianchi Bonomi Hemophilia and Thrombosis Center, Department of Internal Medicine, Istituto di Ricovero e Cura a Carattere Scientifico-Ospedale Maggiore, University of Milan, 20122 Milan, Italy. marco.cattaneo@unimi.it
authors.forename: Marco
authors.initials: M
authors.lastname: Cattaneo
authors.forename: Maddalena L
authors.initials: ML
authors.lastname: Zighetti
authors.forename: Rossana
authors.initials: R
authors.lastname: Lombardi
authors.forename: Constantino
authors.initials: C
authors.lastname: Martinez
authors.forename: Anna
authors.initials: A
authors.lastname: Lecchi
authors.forename: Pamela B
authors.initials: PB
authors.lastname: Conley
authors.forename: Jerry
authors.initials: J
authors.lastname: Ware
authors.forename: Zaverio M
authors.initials: ZM
authors.lastname: Ruggeri
mesh_terms: D000328:Adult; D000527:Alprostadil / Q000378:metabolism; D001483:Base Sequence; D001792:Blood Platelets / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D000242:Cyclic AMP / Q000378:metabolism; D017931:DNA Primers; D005260:Female; D006470:Hemorrhage / Q000151:congenital* / Q000235:genetics; D006579:Heterozygote; D006801:Humans; D008297:Male; D008565:Membrane Proteins; D008875:Middle Aged; D018048:Receptors, Purinergic P2 / Q000235:genetics / Q000378:metabolism*; D058925:Receptors, Purinergic P2Y12; D015398:Signal Transduction
chemical_list: D017931:DNA Primers; D008565:Membrane Proteins; C548576:P2RY12 protein, human; D018048:Receptors, Purinergic P2; D058925:Receptors, Purinergic P2Y12; D000242:Cyclic AMP; D000527:Alprostadil; D002118:Calcium