Differential expression results for <i>CRISPLD2</i> obtained from publicly available data of two previous microarray studies that investigated the effects of GCs on human ASM cells.

<p>Rank refers to ranking within all differentially expressed genes of each comparison group.</p

<i>CRISPLD2</i> is a GC- and IL1β-responsive gene.

<p>ASM cells were treated with 100<b>A</b>) increased <i>CRISPLD2</i> mRNA expression as measured by qRT-PCR, <b>B</b>) increased <i>CRISPLD2</i> protein expression as measured by immuno-blotting. ASM cells were treated with 5 ng/mL IL1β for 24 h, resulting in <b>C</b>) increased <i>CRISPLD2</i> mRNA expression as measured by qRT-PCR, and <b>D</b>) increased <i>CRISPLD2</i> protein expression as measured by immuno-blotting. <i>CRISPLD2</i> mRNA levels were measured in triplicate. CRISPLD2 protein levels are shown as normalized blot densitometry values, and the error bars are SE values across three independent experiments. * <i>P</i><0.05 (<i>t</i> test).</p

<i>CRISPLD2</i> regulates the response to inflammatory cytokines.

<p><b>A</b>) Effect of <i>CRISPLD2</i>-specific siRNA on <i>CRISPLD2</i> mRNA and protein levels. ASM cells were transfected with <i>CRISPLD2</i>-specific siRNA or non-targeting (NT) siRNA, and 72 h later, <i>CRISPLD2</i> mRNA and protein levels were determined by qRT-PCR (levels normalized to those in control cells transfected with NT siRNA) and immuno-blotting, respectively. The effect of <i>CRISPLD2</i> knockdown on IL1β-induced cytokine expression was assessed by transfecting ASM cells with <i>CRISPLD2</i>-specific or NT siRNA, and 72 h later, treating cells for 24 h with <b>B</b>) 5 ng/mL IL1β, <b>C</b>) 100 nM DEX, or <b>D</b>) 5 ng/mL IL1β and 100 nM DEX. <i>IL6</i> expression was determined by qRT-PCR. Normalized mRNA levels are shown. Experiments were performed in triplicate, and the error bars are SE values for three samples. * <i>P</i><0.05 (<i>t</i> test).</p

Top (Q-value <1E-10) differentially expressed genes.

<p>FPKM  =  fragments per kilobase of transcript per million mapped reads.</p

RNA-Seq profiling of DEX-treated ASM cells.

<p><b>A</b>) Volcano plot of overall gene-based differential expression results for four cell lines treated with DEX vs. left untreated (each dot corresponds to a gene). The y-axis corresponds to the negative log (base 10) of P-values while the x-axis corresponds to the negative log (base 2) of the fold change for difference in expression when cells were stimulated with DEX. There were 316 differentially expressed genes according to an adjusted p-value <0.05 (blue dots). <b>B</b>) Validation of known GC-responsive genes through qRT-PCR. After ASM cells were treated with 1 µM DEX for 18 h, the mRNA levels of indicated genes were measured by qRT-PCR and the folds of change in mRNA induced by DEX were calculated. Each column bar represents an individual cell line. Experiments for each cell line were performed in triplicate, and the error bars are SE values corresponding to a cell line's replicates. The dotted line indicates a fold change of 1. ** <i>P</i><0.005, * <i>P</i><0.05 (t test).</p

Validation of GC responsive genes.

<p>After cells from three individual ASM lines were treated with 1 µM DEX for 18 h, the mRNA levels of indicated genes were measured by qRT-PCR and the folds of change induced by DEX were calculated. Each column bar represents an individual cell line; experiments for each cell line were performed in triplicate. Error bars are SE values corresponding to a cell line's replicates.</p

Primary GWAS Top Results (SNPs with Combined P-values <1e-05).

<p>CLLS = CAMP/LOCCS/LODO/Sepracor. CLLS used genotyped SNP data. CARE and ACRN used HapMap Phase 2 imputed data with Mach ratio of empirically observed dosage variance to the expected dosage variance (Rsq) values indicated.</p

Summary of BDR by genotype of the SNP (rs295137) near <i>SPATS2L</i> with lowest P-value among all subjects in the primary populations.

<p>The TT genotype was associated with higher BDR (median 16.0; inter-quartile range (IQR) = [6.2, 32.4]), than the TC genotype (median 11.2; IQR = [5.2, 22.6]) or the CC genotype (median 10.3; IQR = [4.1, 21.7]).</p

Replication study of top SNPs in two independent populations (SARP and DAG).

*<p>rs295137 and rs295114 include SARP and DAG P-values. Remaining SNPs include SARP only.</p

Region of association near <i>SPATS2L</i> SNPs to BDR.

<p>The x-axis denotes position along Chromosome 2. The y-axis denotes −Log₁₀(P) corresponding to 1000GP imputed data P-values. LD between the SNP with the lowest P-value (rs295137) to each SNP in the plot is denoted in colors and was computed according to 1000GP June 2010 CEU data. Plot was created using LocusZoom <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1002824#pgen.1002824-Pruim1" target="_blank">[44]</a>.</p