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.

P2X7 receptor activation enhances SK3 channels- and cystein cathepsin-dependent cancer cells invasiveness

“…Importantly, P2X 7 R were not expressed in non-cancerous mammary cells (MCF-10A, 184A1 and Human Mammary Epithelial Cell) whereas they were markedly expressed in highly invasive MDA-MB-435s cells. Many polymorphisms have been identified in the P2RX7 gene, some of them being responsible for important functional alterations and associated to many diseases (Cabrini et al, 2005;Shemon et al, 2006;Roger et al, 2010b). We therefore performed P2X 7 R complementary DNA sequencing in MDA-MB-435s cells.…”

Purinergic P2X7 receptor functional genetic polymorphisms are associated with the susceptibility to osteoporosis in Chinese postmenopausal women

“…Secondly, rs2230911 polymorphism, which changes serine to threonine at residue 35 (Ser357 to Thr), also resides in the region coding for the intracellular tail of P2X7R and thus was focused on. Mutation of Thr357Ser was shown to be associated with a partial function loss of P2X7R through affecting channel and pore function [23]. It was reported that rs2230911 was associated with a higher prevalence of OP in Scotland women [27].…”

“…However, we do not see the protective pattern for the rs1718119 polymorphism in our study. Fourthly, mutations of rs7958311 could also alter coding of amino acids (His357 to Arg) and was associated with a partial function loss of P2X7R through affecting channel and pore function [22,23]. Rs7958311 was shown to be related with many disorders, including pulmonary tuberculosis, chronic pain, and anxiety [21,28,29].…”

“…It is located in 12q24 and contains 13 exons spanning a 53-kb region [21]. Several single nucleotide polymorphisms (SNPs) in the P2X7R gene have been described to affect the function of this receptor, including loss of function or gain of function [21][22][23][24]. For example, three P2X7R SNPs with amino acid changes, rs3751143 (1513A > C, Glu496Ala), rs2230911 (1096 C > G, Thr357Ser), and rs7958311 (835G > A, Arg270His), were shown to lead to a loss of receptor function in both channel and pore function [22,23].…”

“…Several single nucleotide polymorphisms (SNPs) in the P2X7R gene have been described to affect the function of this receptor, including loss of function or gain of function [21][22][23][24]. For example, three P2X7R SNPs with amino acid changes, rs3751143 (1513A > C, Glu496Ala), rs2230911 (1096 C > G, Thr357Ser), and rs7958311 (835G > A, Arg270His), were shown to lead to a loss of receptor function in both channel and pore function [22,23]. Rs3751143 and rs2230911 were reported to be associated with a higher prevalence of OP [25][26][27], while rs7958311 was shown to be related with other diseases, including pulmonary tuberculosis, chronic pain, and anxiety [21,28,29].…”