IRAK-M is involved in the pathogenesis of early-onset persistent asthma

Abstract

Asthma is a multifactorial disease influenced by genetic and environmental factors. In the past decade, several loci and >100 genes have been found to be associated with the disease in at least one population. Among these loci, region 12q13-24 has been implicated in asthma etiology in multiple populations, suggesting that it harbors one or more asthma susceptibility genes. We performed linkage and association analyses by transmission/disequilibrium test and case-control analysis in the candidate region 12q13-24, using the Sardinian founder population, in which limited heterogeneity of pathogenetic alleles for monogenic and complex disorders as well as of environmental conditions should facilitate the study of multifactorial traits. We analyzed our cohort, using a cutoff age of 13 years at asthma onset, and detected significant linkage to a portion of 12q13-24. We identified IRAK-M as the gene contributing to the linkage and showed that it is associated with early-onset persistent asthma. We defined protective and predisposing SNP haplotypes and replicated associations in an outbred Italian population. Sequence analysis in patients found mutations, including inactivating lesions, in the IRAK-M coding region. Immunohistochemistry of lung biopsies showed that IRAK-M is highly expressed in epithelial cells. We report that IRAK-M is involved in the pathogenesis of early-onset persistent asthma. IRAK-M, a negative regulator of the Toll-like receptor/IL-1R pathways, is a master regulator of NF- kappa B and inflammation. Our data suggest a mechanistic link between hyperactivation of the innate immune system and chronic airway inflammation and indicate IRAK-M as a potential target for therapeutic intervention against asthma.

Figure 1.

Identification of IRAK-M as an asthma susceptibility gene. A, Multipoint linkage analysis plots of chromosome 12q13-24 for Sardinian asthmatic sib pairs. LOD scores are shown for the whole sample (circles), for affected sibs concordant for early age at onset (⩽13 years) (squares), and for affected sibs with at least one patient with age at onset >13 years (triangles). B, Genomic region of the 95% CI for gene location estimate. Asthma candidate genes for which mutation analysis was performed are indicated, along with microsatellite markers used in linkage analysis. C, LD in a 400-kb region containing IRAK-M (black bar). D′ values for pairwise LD between each marker are shown according to LD strength, from D^′>0.8 (red) to D^′<0.3 (white). SNPs used in TDT analysis are also indicated; seven SNPs in bold were significant after multiple test correction (detailed results in table A2).

Figure 2.

Linkage analysis according to the presence of the risk haplotype. With the use of markers from 12q13.3 to 12q21.1 to analyze the initial sample of 100 families (121 sib pairs), the LOD score values relative to 64 sib pairs sharing the risk haplotype (triangles) are similar (MLS 1.70) to those of the whole sample (squares) (MLS 1.93), whereas the sample of siblings not sharing the risk haplotype (circles) shows a much lower value (MLS 0.30) for the same microsatellite marker, D12S75.

Figure 3.

A, B, and C, Immunohistochemical localization of IRAK-M in normal lung biopsy samples from healthy donors. Expression of IRAK-M (A) was detected in the cytoplasm of alveolar macrophages (red arrowhead) and type II epithelial cells, particularly at the level of bronchi (b) and alveoli (a [yellow arrowheads]). Nuclei of epithelial type II cells in bronchi and alveoli are stained by the anti-TTF1 antibody (B) and by the anti-phospho-p65 antibody (C) that recognizes the activated form of the NF-κB subunit. Tissues were counterstained with hematoxylin. Results are representative of at least six independent experiments. a = alveolus; b = bronchus. Yellow arrowhead indicates alveolar type II pneumocytes. Scale bar = 50 μm. D, IRAK-M as a brake on inflammatory processes involved in asthma. Recognition of different elicitors by TLRs on alveolar macrophages and lung epithelial cells triggers the formation of a receptor/adaptor complex including IRAK-1, IRAK-4, and MyD88. Release of phosphorylated IRAK-1 from the signaling complex leads to activation of MAPKs and NF-κB. This in turn results in the production of inflammatory cytokines that can influence T-cell activation and differentiation of Th precursors into Th1 and Th2 subtypes. Once the immune response is mounted, IRAK-M inhibits TLR signaling by interrupting IRAK-1 downstream signal transduction, thus restoring homeostasis. When IRAK-M function is hampered, excessive TLR stimulation may lead to continuous activation of Th1 and Th2 cells in the lung and to the development of atopic asthma and/or asthma exacerbation in response to infections/allergens.