RNAseq analysis of bronchial epithelial cells to identify COPD-associated genes and SNPs

Jiyoun Yeo, Diego A. Morales, Tian Chen, Erin L. Crawford, Xiaolu Zhang, Thomas M. Blomquist, Albert M. Levin, Pierre P. Massion, Douglas A. Arenberg, David Eric Midthun, Peter J. Mazzone, Steven D. Nathan, Ronald J. Wainz, Patrick Nana-Sinkam, Paige F.S. Willey, Taylor J. Arend, Karanbir Padda, Shuhao Qiu, Alexei Federov, Dawn Alita R. HernandezJeffrey R. Hammersley, Youngsook Yoon, Fadi Safi, Sadik A. Khuder, James C. Willey

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Abstract

Background: There is a need for more powerful methods to identify low-effect SNPs that contribute to hereditary COPD pathogenesis. We hypothesized that SNPs contributing to COPD risk through cis-regulatory effects are enriched in genes comprised by bronchial epithelial cell (BEC) expression patterns associated with COPD. Methods: To test this hypothesis, normal BEC specimens were obtained by bronchoscopy from 60 subjects: 30 subjects with COPD defined by spirometry (FEV1/FVC < 0.7, FEV1% < 80%), and 30 non-COPD controls. Targeted next generation sequencing was used to measure total and allele-specific expression of 35 genes in genome maintenance (GM) genes pathways linked to COPD pathogenesis, including seven TP53 and CEBP transcription factor family members. Shrinkage linear discriminant analysis (SLDA) was used to identify COPD-classification models. COPD GWAS were queried for putative cis-regulatory SNPs in the targeted genes. Results: On a network basis, TP53 and CEBP transcription factor pathway gene pair network connections, including key DNA repair gene ERCC5, were significantly different in COPD subjects (e.g., Wilcoxon rank sum test for closeness, p-value = 5.0E-11). ERCC5 SNP rs4150275 association with chronic bronchitis was identified in a set of Lung Health Study (LHS) COPD GWAS SNPs restricted to those in putative regulatory regions within the targeted genes, and this association was validated in the COPDgene non-hispanic white (NHW) GWAS. ERCC5 SNP rs4150275 is linked (D' = 1) to ERCC5 SNP rs17655 which displayed differential allelic expression (DAE) in BEC and is an expression quantitative trait locus (eQTL) in lung tissue (p = 3.2E-7). SNPs in linkage (D' = 1) with rs17655 were predicted to alter miRNA binding (rs873601). A classifier model that comprised gene features CAT, CEBPG, GPX1, KEAP1, TP73, and XPA had pooled 10-fold cross-validation receiver operator characteristic area under the curve of 75.4% (95% CI: 66.3%-89.3%). The prevalence of DAE was higher than expected (p = 0.0023) in the classifier genes. Conclusions: GM genes comprised by COPD-associated BEC expression patterns were enriched for SNPs with cis-regulatory function, including a putative cis-rSNP in ERCC5 that was associated with COPD risk. These findings support additional total and allele-specific expression analysis of gene pathways with high prior likelihood for involvement in COPD pathogenesis.

Original languageEnglish (US)
Article number42
JournalBMC Pulmonary Medicine
Volume18
Issue number1
DOIs
StatePublished - Mar 5 2018

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Chronic Obstructive Pulmonary Disease
Single Nucleotide Polymorphism
Epithelial Cells
Genes
Genome-Wide Association Study
Nonparametric Statistics
Transcription Factors
Alleles
Maintenance
Genome
Gene Expression
Lung
Gene Regulatory Networks
Nucleic Acid Regulatory Sequences
Quantitative Trait Loci
Spirometry
Chronic Bronchitis
Bronchoscopy
Discriminant Analysis
MicroRNAs

Keywords

  • Bronchial epithelial cells
  • CAT
  • CEBPG
  • cis-regulation
  • COPD
  • EQTL
  • ERCC5
  • GPX1
  • GWAS
  • KEAP1
  • TP73
  • XPA

ASJC Scopus subject areas

  • Pulmonary and Respiratory Medicine

Cite this

Yeo, J., Morales, D. A., Chen, T., Crawford, E. L., Zhang, X., Blomquist, T. M., ... Willey, J. C. (2018). RNAseq analysis of bronchial epithelial cells to identify COPD-associated genes and SNPs. BMC Pulmonary Medicine, 18(1), [42]. https://doi.org/10.1186/s12890-018-0603-y

RNAseq analysis of bronchial epithelial cells to identify COPD-associated genes and SNPs. / Yeo, Jiyoun; Morales, Diego A.; Chen, Tian; Crawford, Erin L.; Zhang, Xiaolu; Blomquist, Thomas M.; Levin, Albert M.; Massion, Pierre P.; Arenberg, Douglas A.; Midthun, David Eric; Mazzone, Peter J.; Nathan, Steven D.; Wainz, Ronald J.; Nana-Sinkam, Patrick; Willey, Paige F.S.; Arend, Taylor J.; Padda, Karanbir; Qiu, Shuhao; Federov, Alexei; Hernandez, Dawn Alita R.; Hammersley, Jeffrey R.; Yoon, Youngsook; Safi, Fadi; Khuder, Sadik A.; Willey, James C.

In: BMC Pulmonary Medicine, Vol. 18, No. 1, 42, 05.03.2018.

Research output: Contribution to journalArticle

Yeo, J, Morales, DA, Chen, T, Crawford, EL, Zhang, X, Blomquist, TM, Levin, AM, Massion, PP, Arenberg, DA, Midthun, DE, Mazzone, PJ, Nathan, SD, Wainz, RJ, Nana-Sinkam, P, Willey, PFS, Arend, TJ, Padda, K, Qiu, S, Federov, A, Hernandez, DAR, Hammersley, JR, Yoon, Y, Safi, F, Khuder, SA & Willey, JC 2018, 'RNAseq analysis of bronchial epithelial cells to identify COPD-associated genes and SNPs', BMC Pulmonary Medicine, vol. 18, no. 1, 42. https://doi.org/10.1186/s12890-018-0603-y
Yeo, Jiyoun ; Morales, Diego A. ; Chen, Tian ; Crawford, Erin L. ; Zhang, Xiaolu ; Blomquist, Thomas M. ; Levin, Albert M. ; Massion, Pierre P. ; Arenberg, Douglas A. ; Midthun, David Eric ; Mazzone, Peter J. ; Nathan, Steven D. ; Wainz, Ronald J. ; Nana-Sinkam, Patrick ; Willey, Paige F.S. ; Arend, Taylor J. ; Padda, Karanbir ; Qiu, Shuhao ; Federov, Alexei ; Hernandez, Dawn Alita R. ; Hammersley, Jeffrey R. ; Yoon, Youngsook ; Safi, Fadi ; Khuder, Sadik A. ; Willey, James C. / RNAseq analysis of bronchial epithelial cells to identify COPD-associated genes and SNPs. In: BMC Pulmonary Medicine. 2018 ; Vol. 18, No. 1.
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abstract = "Background: There is a need for more powerful methods to identify low-effect SNPs that contribute to hereditary COPD pathogenesis. We hypothesized that SNPs contributing to COPD risk through cis-regulatory effects are enriched in genes comprised by bronchial epithelial cell (BEC) expression patterns associated with COPD. Methods: To test this hypothesis, normal BEC specimens were obtained by bronchoscopy from 60 subjects: 30 subjects with COPD defined by spirometry (FEV1/FVC < 0.7, FEV1{\%} < 80{\%}), and 30 non-COPD controls. Targeted next generation sequencing was used to measure total and allele-specific expression of 35 genes in genome maintenance (GM) genes pathways linked to COPD pathogenesis, including seven TP53 and CEBP transcription factor family members. Shrinkage linear discriminant analysis (SLDA) was used to identify COPD-classification models. COPD GWAS were queried for putative cis-regulatory SNPs in the targeted genes. Results: On a network basis, TP53 and CEBP transcription factor pathway gene pair network connections, including key DNA repair gene ERCC5, were significantly different in COPD subjects (e.g., Wilcoxon rank sum test for closeness, p-value = 5.0E-11). ERCC5 SNP rs4150275 association with chronic bronchitis was identified in a set of Lung Health Study (LHS) COPD GWAS SNPs restricted to those in putative regulatory regions within the targeted genes, and this association was validated in the COPDgene non-hispanic white (NHW) GWAS. ERCC5 SNP rs4150275 is linked (D' = 1) to ERCC5 SNP rs17655 which displayed differential allelic expression (DAE) in BEC and is an expression quantitative trait locus (eQTL) in lung tissue (p = 3.2E-7). SNPs in linkage (D' = 1) with rs17655 were predicted to alter miRNA binding (rs873601). A classifier model that comprised gene features CAT, CEBPG, GPX1, KEAP1, TP73, and XPA had pooled 10-fold cross-validation receiver operator characteristic area under the curve of 75.4{\%} (95{\%} CI: 66.3{\%}-89.3{\%}). The prevalence of DAE was higher than expected (p = 0.0023) in the classifier genes. Conclusions: GM genes comprised by COPD-associated BEC expression patterns were enriched for SNPs with cis-regulatory function, including a putative cis-rSNP in ERCC5 that was associated with COPD risk. These findings support additional total and allele-specific expression analysis of gene pathways with high prior likelihood for involvement in COPD pathogenesis.",
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TY - JOUR

T1 - RNAseq analysis of bronchial epithelial cells to identify COPD-associated genes and SNPs

AU - Yeo, Jiyoun

AU - Morales, Diego A.

AU - Chen, Tian

AU - Crawford, Erin L.

AU - Zhang, Xiaolu

AU - Blomquist, Thomas M.

AU - Levin, Albert M.

AU - Massion, Pierre P.

AU - Arenberg, Douglas A.

AU - Midthun, David Eric

AU - Mazzone, Peter J.

AU - Nathan, Steven D.

AU - Wainz, Ronald J.

AU - Nana-Sinkam, Patrick

AU - Willey, Paige F.S.

AU - Arend, Taylor J.

AU - Padda, Karanbir

AU - Qiu, Shuhao

AU - Federov, Alexei

AU - Hernandez, Dawn Alita R.

AU - Hammersley, Jeffrey R.

AU - Yoon, Youngsook

AU - Safi, Fadi

AU - Khuder, Sadik A.

AU - Willey, James C.

PY - 2018/3/5

Y1 - 2018/3/5

N2 - Background: There is a need for more powerful methods to identify low-effect SNPs that contribute to hereditary COPD pathogenesis. We hypothesized that SNPs contributing to COPD risk through cis-regulatory effects are enriched in genes comprised by bronchial epithelial cell (BEC) expression patterns associated with COPD. Methods: To test this hypothesis, normal BEC specimens were obtained by bronchoscopy from 60 subjects: 30 subjects with COPD defined by spirometry (FEV1/FVC < 0.7, FEV1% < 80%), and 30 non-COPD controls. Targeted next generation sequencing was used to measure total and allele-specific expression of 35 genes in genome maintenance (GM) genes pathways linked to COPD pathogenesis, including seven TP53 and CEBP transcription factor family members. Shrinkage linear discriminant analysis (SLDA) was used to identify COPD-classification models. COPD GWAS were queried for putative cis-regulatory SNPs in the targeted genes. Results: On a network basis, TP53 and CEBP transcription factor pathway gene pair network connections, including key DNA repair gene ERCC5, were significantly different in COPD subjects (e.g., Wilcoxon rank sum test for closeness, p-value = 5.0E-11). ERCC5 SNP rs4150275 association with chronic bronchitis was identified in a set of Lung Health Study (LHS) COPD GWAS SNPs restricted to those in putative regulatory regions within the targeted genes, and this association was validated in the COPDgene non-hispanic white (NHW) GWAS. ERCC5 SNP rs4150275 is linked (D' = 1) to ERCC5 SNP rs17655 which displayed differential allelic expression (DAE) in BEC and is an expression quantitative trait locus (eQTL) in lung tissue (p = 3.2E-7). SNPs in linkage (D' = 1) with rs17655 were predicted to alter miRNA binding (rs873601). A classifier model that comprised gene features CAT, CEBPG, GPX1, KEAP1, TP73, and XPA had pooled 10-fold cross-validation receiver operator characteristic area under the curve of 75.4% (95% CI: 66.3%-89.3%). The prevalence of DAE was higher than expected (p = 0.0023) in the classifier genes. Conclusions: GM genes comprised by COPD-associated BEC expression patterns were enriched for SNPs with cis-regulatory function, including a putative cis-rSNP in ERCC5 that was associated with COPD risk. These findings support additional total and allele-specific expression analysis of gene pathways with high prior likelihood for involvement in COPD pathogenesis.

AB - Background: There is a need for more powerful methods to identify low-effect SNPs that contribute to hereditary COPD pathogenesis. We hypothesized that SNPs contributing to COPD risk through cis-regulatory effects are enriched in genes comprised by bronchial epithelial cell (BEC) expression patterns associated with COPD. Methods: To test this hypothesis, normal BEC specimens were obtained by bronchoscopy from 60 subjects: 30 subjects with COPD defined by spirometry (FEV1/FVC < 0.7, FEV1% < 80%), and 30 non-COPD controls. Targeted next generation sequencing was used to measure total and allele-specific expression of 35 genes in genome maintenance (GM) genes pathways linked to COPD pathogenesis, including seven TP53 and CEBP transcription factor family members. Shrinkage linear discriminant analysis (SLDA) was used to identify COPD-classification models. COPD GWAS were queried for putative cis-regulatory SNPs in the targeted genes. Results: On a network basis, TP53 and CEBP transcription factor pathway gene pair network connections, including key DNA repair gene ERCC5, were significantly different in COPD subjects (e.g., Wilcoxon rank sum test for closeness, p-value = 5.0E-11). ERCC5 SNP rs4150275 association with chronic bronchitis was identified in a set of Lung Health Study (LHS) COPD GWAS SNPs restricted to those in putative regulatory regions within the targeted genes, and this association was validated in the COPDgene non-hispanic white (NHW) GWAS. ERCC5 SNP rs4150275 is linked (D' = 1) to ERCC5 SNP rs17655 which displayed differential allelic expression (DAE) in BEC and is an expression quantitative trait locus (eQTL) in lung tissue (p = 3.2E-7). SNPs in linkage (D' = 1) with rs17655 were predicted to alter miRNA binding (rs873601). A classifier model that comprised gene features CAT, CEBPG, GPX1, KEAP1, TP73, and XPA had pooled 10-fold cross-validation receiver operator characteristic area under the curve of 75.4% (95% CI: 66.3%-89.3%). The prevalence of DAE was higher than expected (p = 0.0023) in the classifier genes. Conclusions: GM genes comprised by COPD-associated BEC expression patterns were enriched for SNPs with cis-regulatory function, including a putative cis-rSNP in ERCC5 that was associated with COPD risk. These findings support additional total and allele-specific expression analysis of gene pathways with high prior likelihood for involvement in COPD pathogenesis.

KW - Bronchial epithelial cells

KW - CAT

KW - CEBPG

KW - cis-regulation

KW - COPD

KW - EQTL

KW - ERCC5

KW - GPX1

KW - GWAS

KW - KEAP1

KW - TP73

KW - XPA

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U2 - 10.1186/s12890-018-0603-y

DO - 10.1186/s12890-018-0603-y

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