title: A Thr357 to Ser polymorphism in homozygous and compound heterozygous subjects causes absent or reduced P2X7 function and impairs ATP-induced mycobacterial killing by macrophages.
abstract: The P2X(7) receptor is a ligand-gated cation channel that is highly expressed on mononuclear leukocytes and that mediates ATP-induced apoptosis and killing of intracellular pathogens. There is a wide variation in P2X(7) receptor function between subjects, explained in part by four loss-of-function polymorphisms (R307Q, E496A, I568N, and a 5'-intronic splice site polymorphism), as well as rare mutations. In this study, we report the allele frequencies of 11 non-synonymous P2X(7) polymorphisms and describe a fifth loss-of-function polymorphism in the gene (1096C --> G), which changes Thr(357) to Ser (T357S) with an allele frequency of 0.08 in the Caucasian population. P2X(7) function was measured by ATP-induced ethidium(+) influx into peripheral blood lymphocytes and monocytes and, when compared with wild-type subjects, was reduced to 10-65% in heterozygotes, 1-18% in homozygotes, and 0-10% in compound heterozygotes carrying T357S and a second loss-of-function polymorphism. Overexpression of the T357S mutant P2X(7) in either HEK-293 cells or Xenopus oocytes gave P2X(7) function of approximately 50% that of wild-type constructs. Differentiation of monocytes to macrophages, which also up-regulates P2X(7), restored P2X(7) function to near normal in cells heterozygous for T357S and to a value 50-65% of wild-type in cells homozygous for T357S or compound heterozygous for T357S/E496A. However, macrophages from subjects that are compound heterozygous for either T357S/R307Q or T357S/stop codon had near-to-absent P2X(7) function. These functional deficits induced by T357S were paralleled by impaired ATP-induced apoptosis and mycobacteria killing in macrophages from these subjects. Lymphocytes, monocytes, and macrophages from subjects homozygous for T357S or compound heterozygous for T357S and a second loss-of-function allele have reduced or absent P2X(7) receptor function.
authors.affiliation: Department of Medicine, University of Sydney, Level 5, Spurrett Building, Nepean Hospital, Penrith, New South Wales 2750, Australia.
authors.forename: Anne N
authors.initials: AN
authors.lastname: Shemon
authors.forename: Ronald
authors.initials: R
authors.lastname: Sluyter
authors.forename: Suran L
authors.initials: SL
authors.lastname: Fernando
authors.forename: Alison L
authors.initials: AL
authors.lastname: Clarke
authors.forename: Lan-Phuong
authors.initials: LP
authors.lastname: Dao-Ung
authors.forename: Kristen K
authors.initials: KK
authors.lastname: Skarratt
authors.forename: Bernadette M
authors.initials: BM
authors.lastname: Saunders
authors.forename: Khai See
authors.initials: KS
authors.lastname: Tan
authors.forename: Ben J
authors.initials: BJ
authors.lastname: Gu
authors.forename: Stephen J
authors.initials: SJ
authors.lastname: Fuller
authors.forename: Warwick J
authors.initials: WJ
authors.lastname: Britton
authors.forename: Steven
authors.initials: S
authors.lastname: Petrou
authors.forename: James S
authors.initials: JS
authors.lastname: Wiley
mesh_terms: D000255:Adenosine Triphosphate / Q000737:chemistry*; D000483:Alleles; D000818:Animals; D001464:Barium / Q000494:pharmacology; D002454:Cell Differentiation; D002460:Cell Line; D018388:Codon, Terminator; D004594:Electrophysiology; D004996:Ethidium / Q000494:pharmacology; D005260:Female; D005434:Flow Cytometry; D005787:Gene Frequency; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006579:Heterozygote; D006720:Homozygote; D006801:Humans; D007371:Interferon-gamma / Q000378:metabolism; D008214:Lymphocytes / Q000166:cytology / Q000378:metabolism / Q000382:microbiology; D008264:Macrophages / Q000378:metabolism* / Q000382:microbiology; D008297:Male; D008856:Microscopy, Fluorescence; D009000:Monocytes / Q000166:cytology / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D009161:Mycobacterium / Q000378:metabolism*; D009163:Mycobacterium bovis / Q000378:metabolism; D009865:Oocytes / Q000378:metabolism; D011110:Polymorphism, Genetic; D020641:Polymorphism, Single Nucleotide; D018048:Receptors, Purinergic P2 / Q000235:genetics*; D058486:Receptors, Purinergic P2X7; D012694:Serine / Q000737:chemistry / Q000235:genetics*; D013912:Threonine / Q000737:chemistry / Q000235:genetics*; D014162:Transfection; D014982:Xenopus laevis
chemical_list: D018388:Codon, Terminator; C546468:P2RX7 protein, human; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; D049452:Green Fluorescent Proteins; D001464:Barium; D013912:Threonine; D012694:Serine; D007371:Interferon-gamma; D000255:Adenosine Triphosphate; D004996:Ethidium