Functional Variant in the Autophagy-Related 5 Gene Promotor is Associated with Childhood Asthma

Rationale and Objective: Autophagy is a cellular process directed at eliminating or recycling cellular proteins. Recently, the autophagy pathway has been implicated in immune dysfunction, the pathogenesis of inflammatory disorders, and response to viral infection. Associations between two genes in the autophagy pathway, ATG5 and ATG7, with childhood asthma were investigated. Methods: Using genetic and experimental approaches, we examined the association of 13 HapMap-derived tagging SNPs in ATG5 and ATG7 with childhood asthma in 312 asthmatic and 246 non-allergic control children. We confirmed our findings by using independent cohorts and imputation analysis. Finally, we evaluated the functional relevance of a disease associated SNP. Measurements and Main Results: We demonstrated that ATG5 single nucleotide polymorphisms rs12201458 and rs510432 were associated with asthma (p = 0.00085 and 0.0025, respectively). In three independent cohorts, additional variants in ATG5 in the same LD block were associated with asthma (p,0.05). We found that rs510432 was functionally relevant and conferred significantly increased promotor activity. Furthermore, Atg5 expression was increased in nasal epithelium of acute asthmatics compared to stable asthmatics and non-asthmatic controls. Conclusion: Genetic variants in ATG5, including a functional promotor variant, are associated with childhood asthma. Thes

The E3 ubiquitin ligase STUB1 regulates autophagy and mitochondrial biogenesis by modulating TFEB activity

TFEB is a master regulator for transcription of genes involved in autophagy, lysosome and mitochondrial biogenesis. Activity of TFEB is inhibited upon its phosphorylation. STUB1, a chaperone-dependent E3 ubiquitin ligase, modulates TFEB activity by preferentially targeting inactive phosphorylated TFEB for degradation by the ubiquitin proteasome pathway. Thus, the ubiquitin-proteasome pathway participates in regulating autophagy and lysosomal functions by regulating the activity of TFEB

STUB1 regulates TFEB-induced autophagy-lysosome pathway

TFEB is a master regulator for transcription of genes involved in autophagy and lysosome biogenesis. Activity of TFEB is inhibited upon its serine phosphorylation by mTOR The overall mechanisms by which TFEB activity in the cell is regulated are not well elucidated. Specifically, the mechanisms of TFEB turnover and how they might influence its activity remain unknown. Here, we show that STUB1, a chaperone-dependent E3 ubiquitin ligase, modulates TFEB activity by preferentially targeting inactive phosphorylated TFEB for degradation by the ubiquitin-proteasome pathway. Phosphorylated TFEB accumulated in STUB1-deficient cells and in tissues of STUB1-deficient mice resulting in reduced TFEB activity. Conversely, cellular overexpression of STUB1 resulted in reduced phosphorylated TFEB and increased TFEB activity. STUB1 preferentially interacted with and ubiqutinated phosphorylated TFEB, targeting it to proteasomal degradation. Consistent with reduced TFEB activity, accumulation of phosphorylated TFEB in STUB1-deficient cells resulted in reduced autophagy and reduced mitochondrial biogenesis. These studies reveal that the ubiquitin-proteasome pathway participates in regulating autophagy and lysosomal functions by regulating the activity of TFEB

Characteristics of the GCPCR population.

a<p>Indicates sample passing quality control.</p>b<p>Indicates significant differences (p<0.05) with non-allergic control children.</p

Genotyped SNPs and their minor allele frequencies (MAF) in the GCPCR cohort.

a<p>p = 0.00085; OR = 0.52, 95% CI, 0.36–0.77.</p>b<p>p = 0.0025; OR = 1.47, 95% CI, 1.14–1.88.</p

Asthma-associated rs510432 SNP G variant allele confers enhanced promotor activity.

<p>A. ATG5 promotor fragments generated from genomic DNA isolated from human peripheral blood mononuclear cells. B, C. Luciferase activity of promotor assay vectors. In the mutant the corresponding sites of the two strands of the DNA were mutated from A to G. Promotor activities (corrected for transfection efficiency) are presented as fold increase relative to empty vector (PGL4.20). The fold ratio of empty PGL4.20 Firefly Luciferase plasmid to PGL4.73 Renilla transfection control vector was normalized to 1. Mean ± SD, n = 3 independent experiments.</p