The neuronal ␣7 nicotinic receptor subunit gene (CHRNA7) is partially duplicated in the human genome forming a hybrid gene (CHRFAM7A) with the novel FAM7A gene. The hybrid gene transcript, dup␣7, has been identified in brain, immune cells, and the HL-60 cell line, although its translation and function are still unknown. In this study, dup␣7 cDNA has been cloned and expressed in GH4C1 cells and Xenopus oocytes to study the pattern and functional role of the expressed protein. Our results reveal that dup␣7 transcript was natively translated in HL-60 cells and heterologously expressed in GH4C1 cells and oocytes. Injection of dup␣7 mRNA into oocytes failed to generate functional receptors, but when co-injected with ␣7 mRNA at ␣7/dup␣7 ratios of 5:1, 2:1, 1:1, 1:5, and 1:10, it reduced the nicotine-elicited ␣7 current generated in control oocytes (␣7 alone) by 26, 53, 75, 93, and 94%, respectively. This effect is mainly due to a reduction in the number of functional ␣7 receptors reaching the oocyte membrane, as deduced from ␣-bungarotoxin binding and fluorescent confocal assays. Two additional findings open the possibility that the dominant negative effect of dup␣7 on ␣7 receptor activity observed in vitro could be extrapolated to in vivo situations. (i) Compared with ␣7 mRNA, basal dup␣7 mRNA levels are substantial in human cerebral cortex and higher in macrophages.(ii) dup␣7 mRNA levels in macrophages are down-regulated by IL-1␤, LPS, and nicotine. Thus, dup␣7 could modulate ␣7 receptor-mediated synaptic transmission and cholinergic antiinflammatory response.Neuronal ␣7 nicotinic acetylcholine receptors (␣7 nAChRs) 4 are widely expressed in the central and peripheral nervous systems. In neurons, homomeric ␣7 nAChRs, composed of five ␣7 subunits, modulate neurotransmitter release in presynaptic nerve terminals and induce excitatory impulses in postsynaptic neurons (1-4). Signaling through ␣7 nAChRs in the central nervous system has been associated with neuronal plasticity and cell survival (5-7), although impaired activity of this receptor has been implicated in the pathogenesis of schizophrenia, Alzheimer disease, and depression (8 -12). The presence of ␣7 nAChRs has also been reported in non-neuronal cells such as vascular and brain en-

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Nicotine stimulation of α7 nicotinic acetylcholine receptor (α7 nAChR) powerfully inhibits pro-inflammatory cytokine production in lipopolysaccharide (LPS)-stimulated macrophages and in experimental models of endotoxemia. A signaling pathway downstream from the α7 nAChRs, which involves the collaboration of JAK2/STAT3 and NF-κB to interfere with signaling by Toll-like receptors (TLRs), has been implicated in this anti-inflammatory effect of nicotine. Here, we identifiy an alternative mechanism involving interleukin-1 receptor-associated kinase M (IRAK-M), a negative regulator of innate TLR-mediated immune responses. Our data show that nicotine up-regulates IRAK-M expression at the mRNA and protein level in human macrophages, and that this effect is secondary to α7 nAChR activation. By using selective inhibitors of different signaling molecules downstream from the receptor, we provide evidence that activation of STAT3, via either JAK2 and/or PI3K, through a single (JAK2/PI3K/STAT3) or two convergent cascades (JAK2/STAT3 and PI3K/STAT3), is necessary for nicotine-induced IRAK-M expression. Moreover, down-regulation of this expression by small interfering RNAs specific to the IRAK-M gene significantly reverses the anti-inflammatory effect of nicotine on LPS-induced TNF-α production. Interestingly, macrophages pre-exposed to nicotine exhibit higher IRAK-M levels and reduced TNF-α response to an additional LPS challenge, a behavior reminiscent of the ‘endotoxin tolerant’ phenotype identified in monocytes either pre-exposed to LPS or from immunocompromised septic patients. Since nicotine is a major component of tobacco smoke and increased IRAK-M expression has been considered one of the molecular determinants for the induction of the tolerant phenotype, our findings showing IRAK-M overexpression could partially explain the known influence of smoking on the onset and progression of inflammatory and infectious diseases.

This study reveals that the PBMC α7 gene expression level is a clinically relevant marker for CAP activity in sepsis: the higher the α7 expression, the better the inflammation control and the prognosis.

Cigarette smoking is associated with increased risk for all histologic types of lung cancer, but why the strength of this association is stronger for squamous cell carcinoma than adenocarcinoma of the lung (SQC-L, ADC-L) is not fully understood. Because nicotine and tobacco-specific nitrosamines contribute to carcinogenesis by activating nicotinic acetylcholine receptors (nAChRs) on lung tumors and epithelial cells, we investigated whether differential expression of nAChR subtypes in these tumors could explain their different association with smoking. Expression of nAChR subunit genes in paired tumor and non-tumor lung specimens from 40 SQC-L and 38 ADC-L patients was analyzed by quantitative PCR. Compared to normal lung, both tumors share: i) transcriptional dysregulation of CHRNA3/CHRNA5/CHRNB4 (α3, α5, β4 subunits) at the chromosomal locus that predisposes to lung cancer; and ii) decreased expression of CHRFAM7A (dupα7 subunit); this last subunit negatively modulates α7-nAChR activity in oocytes. In contrast, CHRNA7 (α7 subunit) expression was increased in SQC-L, particularly in smokers and non-survivors, while CHRNA4 (α4 subunit) expression was decreased in ADC-L. Thus, over-representation of cancer-stimulating α7-nAChR in SQC-L, also potentiated by smoking, and under-representation of cancer-inhibiting α4β2-nAChR in ADC-L could explain the different tobacco influences on the tumorigenic process in each cancer type.