Our data support a role for the autophagy pathway, and specifically ATG5 but not ATG7, in childhood asthma. We identified two variants in the ATG5 gene that are associated with asthma using a genetic association study and uncovered additional novel associations using genotypes inferred through imputation. We investigated the biologic relevance of the ATG5 rs510432 SNP and found that the disease-associated allelic variant confers enhanced ATG5 promotor activity. When we queried the TRANSFAC database for transcription factor binding sites that include rs510432, we identified two transcription factors, STAT1 and C-Fos, which have been associated with asthma and may be mediating the functional differences observed. Further, we demonstrated that Atg5 mRNA expression is up-regulated in human nasal epithelial cells during an acute asthma attack. Collectively, our results suggest a novel function for ATG5 in asthma.
By screening the autophagy genes ATG5
, we found evidence of association with variants in ATG5
with childhood asthma. This is a novel finding as there have not been any reported associations of ATG5
as a candidate gene for asthma based on searches using PUBMED (terms ATG5
genetic association, date 8/29/2011) and the National Human Genome Research Institute’s Catalog of Published Genome-Wide Association Studies (http://www.genome.gov/gwastudies/index.cfm?pageid=26525384#searchForm
assessed 8/29/2011) 
. One of our primary findings was that the promotor variant rs510432 was associated with childhood asthma in the GCPCR cohort, where asthmatics had a 1.47 fold increase of having the minor allele G compared to controls. To determine whether this promotor variant had functional effects, the promotor activity of ATG5
variants with the A and G alleles were compared. The G allele had higher promotor activity than the A allele. The increased promotor activity of the allele associated with increased risk of asthma (G) is consistent with our gene expression studies showing increased gene expression of ATG5
in asthmatics. Further, the increased promotor activity is consistent with the previous study demonstrating increased activation of autophagy proteins in lung tissue from chronic obstructive pulmonary disease patients 
Several other ATG5
SNPs were found to be associated with childhood asthma. The genotyped ATG5
rs12201458 SNP is located in intron 7, the last intron in the gene. This SNP is predicted to be an intronic enhancer with a low predicted risk of functional effects using FASTSNP 
. As, the SNPs genotyped for ATG5
were predominantly tagging, the identified association with rs122201458 may be due to linkage disequilibrium with functional variants. Thus, the genotypes at markers that had not been genotyped in this study were imputed using a reference panel. Imputation can permit the comparison of studies which focused on different SNPs. Using meta-analysis, four SNPs exhibited significant association across the studies.
We found Atg5
expression to be up-regulated in nasal epithelium from acute asthmatics. While there have been no previous studies investigating Atg5
expression in asthmatics, there is evidence of increased expression of autophagic proteins, including Atg5
, in lung tissue from patients with chronic obstructive pulmonary disease 
. The respiratory tract is divided into the upper airway (UA; the portion from the nose to the vocal cords) and the lower airway (LA; below the vocal cords). Epidemiologic and biologic evidence support this concept of a “united airway” in which the UA reflects pathophysiologic changes occurring in the LA and vice versa through biological cross-talk. Studies have demonstrated that comparable inflammatory processes underlie rhinitis and asthma 
; and not only does nasal allergen challenge initiate pulmonary inflammation 
, but lung allergen challenge induces inflammation in bronchial and nasal mucosa 
. The autophagy pathway is responsive to cigarette smoke exposure 
and viral infection 
, important cofactors for asthma, further supporting a role of autophagy in respiratory disease.
is necessary for antigen presentation 
and can lead to increased viral clearance 
, autophagy machinery can also be hijacked to increase viral replication 
, though indispensible for autophagy, has functions independent of autophagy including a role in apoptosis and regulation of interferon (IFN) responses against viral infections 
. Indeed, the Atg12-Atg5
conjugate has been shown to negatively regulate the type I IFN modulating pathway 
. Thus, in contrast to anti-pathogenic properties of autophagic processes, Atg5
also has the capacity to promote RNA virus replication by inhibiting innate anti-virus immune responses, a rather paradoxical role for Atg5
. These non-canonical roles for Atg5
in regulation of apoptosis and IFN production could have significance in asthma pathology in relation to immune responses to viral infections. Our data has demonstrated amplified Atg5
expression in acute asthmatics. Consistent with previous studies indicating that asthmatics have slower viral clearance 
, increased Atg5
expression could lead to augmented viral replication, greater virus production and thus prolonged viral clearance. Taken together, these studies provide a potential mechanism for a role of ATG5 in asthma. Future studies are required to determine if ATG5 has a causal role in asthma or if these differences are due to inflammation and related cell death.
A major strength of our study is the use of functional investigations to complement the genetic associations. Indeed, many genetic association studies have reported associations with asthma, but few have characterized functional effects 
. Our data provide strong evidence for association of a functional promotor SNP in ATG5
with childhood asthma. Our study is limited in that we utilized adult controls in one of the cohorts. Although many studies prefer to match cases and controls on age, studies have utilized adults as controls for childhood asthma as they represent a truly asthma-free population, whereas similarly aged child controls may go on to develop asthma 
. While this has a potential to create bias, this cohort was used only to confirm findings and therefore the risk of incorrect inference is minimal. Some of the cohorts were of modest size, but it is important to note that both the CAMP and CARE cohorts have extremely well characterized subjects with detailed phenotypic data. However, replication in additional cohorts is warranted.
In summary, several independent investigators have linked autophagy to various aspects of the innate and adaptive immunity. ATG5 is a key component of the autophagy machinery and has functions in viral clearance. We have demonstrated that ATG5 variants are associated with childhood asthma, including a variant that confers enhanced promotor activity and that Atg5 expression is dysregulated in children with asthma. Additional studies are necessary to further elucidate biological roles of autophagy and autophagy-related antiviral defense in asthma pathogenesis.