Transcriptome Profiling of Human Airway Smooth Muscle Cells Stimulated with Dexamethasone Identifies CRISPLD2 as a Regulator of Steroid and Immune Response. B. E. Himes1,2,3, X. Jiang4, P. Wagner4, R. Hu4, B. Klanderman2, Q. Duan1, J. Lasky-Su1, C. Nikolos5, W. Jester5, M. Johnson5, R. A. Panettieri Jr.5, K. G. Tantisira1, S. T. Weiss1,2, Q. Lu4 1) Channing Division of Network Medicine, Brigham and Women's Hospital and Harvard Medical School, Boston, MA; 2) Partners Center for Personalized Genetic Medicine, Boston, MA; 3) Childrens Hospital Informatics Program, Boston, MA; 4) Program in Molecular and Integrative Physiological Sciences, Department of Environmental Health, Harvard School of Public Health, Boston, MA; 5) Pulmonary, Allergy and Critical Care Division, University of Pennsylvania, Philadelphia, PA.
Asthma is a chronic inflammatory airway disease that affects over 300 million people around the world. Glucocorticoids are common medications used to manage inflammatory diseases, and one of the primary tissues that glucocorticoids target in the treatment of asthma is the airway smooth muscle. RNA-Seq is a high-throughput sequencing method that provides comprehensive expression analysis, including discovery of novel genes, non-coding transcripts, and splice variants. We used RNA-Seq to characterize changes in the human airway smooth muscle (HASM) transcriptome in response to treatment with a glucocorticoid (i.e. dexamethasone, 1 M for 18h) using primary HASM cells from four white male donors. The Illumina TruSeq method was used to prepare RNA-seq libraries that were sequenced with an Illumina HiSeq 2000 instrument and aligned to the hg19 reference genome using TopHat. Differential expression analysis, carried out using Cufflinks and CummeRbund, revealed that 316 genes were significantly differentially expressed between control and dex-treated cells. Such genes included ones that have been previously related to steroid responsiveness and inflammation (i.e., DUSP1, KLF15, PER1, and TSC22D3 (a.k.a. GILZ), as well as novel candidates that are not known to be associated with glucocorticoid response. Previously conducted GWAS of two glucocorticoid- and HASM-related phenotypes, inhaled corticosteroid resistance and bronchodilator response, were used to screen differentially expressed genes. One of the top dex-induced genes, CRISPLD2, had SNPs with nominal associations (P-value <10-03) with both of these asthma phenotypes and was selected for further functional studies. After verifying that dexamethasone significantly increases both CRISPLD2 mRNA and protein levels in HASM cells, we performed knock-down experiments in which CRISPLD2 was found to (1) reverse the effect of dexamethasone on two known steroid-response genes (i.e. ENaC and TSC22D3) and (2) increase the IL1 responsiveness of a inflammatory gene (i.e. IL6). Our results suggest that CRISPLD2 regulates both steroid and NFB-dependent immune responses and may play a role in asthma pharmacogenetics.
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