INSERM U1085/IRSET, IFR140, Université de Rennes 1, 2 Avenue du Pr. L. Bernard, Rennes 35043, France.
J Biol Chem. 2012 Feb 3;287(6):4041-52. doi: 10.1074/jbc.M111.319970. Epub 2011 Dec 13.
Polycyclic aromatic hydrocarbons (PAHs) such as benzo(a)pyrene (B(a)P) are widely distributed environmental contaminants, known as potent ligands of the aryl hydrocarbon receptor (AhR). These chemicals trigger an early and transient increase of intracellular calcium concentration (Ca(2+)), required for AhR-related effects of PAHs. The mechanisms involved in this calcium mobilization were investigated in the present study. We demonstrated that B(a)P-mediated Ca(2+) induction was prevented in endothelial HMEC-1 cells by counteracting β2-adrenoreceptor (β2ADR) activity using pharmacological antagonists, anti-β2ADR antibodies, or siRNA-mediated knockdown of β2ADR expression; by contrast, it was strongly potentiated by β2ADR overexpression in human kidney HEK293 cells. B(a)P was shown, moreover, to directly bind to β2ADR, as assessed by in vitro binding assays and molecular modeling. Pharmacological inhibition and/or siRNA-mediated silencing of various signaling actors acting downstream of β2ADR in a sequential manner, such as G protein, adenylyl cyclase, Epac-1 protein, and inositol 1,4,5-trisphosphate (IP(3))/IP(3) receptor, were next demonstrated to prevent B(a)P-induced calcium signal. Inhibition or knockdown of these signaling elements, as well as the use of chemical β-blockers, were finally shown to counteract B(a)P-mediated induction of cytochrome P-450 1B1, a prototypical AhR target gene. Taken together, our results show that B(a)P binds directly to β2ADR and consequently utilizes β2ADR machinery to mobilize Ca(2+), through activation of a G protein/adenylyl cyclase/cAMP/Epac-1/IP(3) pathway. This β2ADR-dependent signaling pathway activated by PAHs may likely be crucial for PAH-mediated up-regulation of AhR target genes, thus suggesting a contribution of β2ADR to the health-threatening effects of these environmental pollutants.
多环芳烃（PAHs）如苯并（a）芘（B（a）P）是广泛分布的环境污染物，被称为芳烃受体（AhR）的有效配体。这些化学物质会触发细胞内钙离子浓度（[Ca（2+）]（i））的早期和短暂增加，这是 PAHs 产生 AhR 相关效应所必需的。本研究探讨了这种钙动员的机制。我们证明，内皮 HMEC-1 细胞中 B（a）P 介导的 [Ca（2+）]（i）诱导可通过使用药理学拮抗剂、抗β2-肾上腺素能受体（β2ADR）抗体或 siRNA 介导的β2ADR 表达敲低来拮抗β2ADR 活性来预防；相比之下，在人肾 HEK293 细胞中，β2ADR 的过表达强烈增强了 B（a）P 的诱导作用。此外，通过体外结合测定和分子建模，我们证明 B（a）P 可直接与β2ADR 结合。接下来，我们依次以顺序方式抑制和/或 siRNA 介导沉默β2ADR 下游的各种信号转导因子，如 G 蛋白、腺苷酸环化酶、Epac-1 蛋白和肌醇 1,4,5-三磷酸（IP（3））/IP（3）受体，结果表明这可预防 B（a）P 诱导的钙信号。最后，我们还证明了这些信号元件的抑制或敲低，以及化学β-阻滞剂的使用，可拮抗 B（a）P 介导的细胞色素 P-450 1B1 的诱导，这是 AhR 靶基因的典型代表。总之，我们的结果表明，B（a）P 直接与β2ADR 结合，然后通过激活 G 蛋白/腺苷酸环化酶/cAMP/Epac-1/IP（3）途径，利用β2ADR 机制来动员 [Ca（2+）]（i）。这种由 PAHs 激活的β2ADR 依赖性信号通路可能对 PAH 介导的 AhR 靶基因的上调至关重要，这表明β2ADR 对这些环境污染物的健康威胁效应有一定的贡献。
