A novel proteogenomics approach identifying key proteins in severe COPD

Rationale COPD is influenced by many genetic and environmental factors. Genetic studies provided insights in the molecular mechanisms underlying COPD, but did not inform on protein changes. We propose that whichever factor contributes to disease, the final common pathway for disease development lies in the proteome. Therefore, we performed a combined proteome and RNA sequencing analysis comparing severe COPD and non-COPD control lung tissue. Methods Frozen lung tissue samples from 10 ex-smoking stage IV COPD patients and 10 ex-smoking non-COPD controls were used for protein and RNA extraction using urea and ammonium bicarbonate buffer and Trizol, respectively. The trypsin digested protein extract was analysed with LC-MS/MS approach using the Q-Exactive Plus (Thermo Scientific). RNA sequencing was performed with polyA-selected RNA sequencing using strand-specific quantitative NextFlex qRNA-Seq kit (Bio scientific) on Illumina HiSeq 2500 equipment with 20 million sequencing reads. We constructed patient specific protein reference databases using the RNA sequencing data and these were used to optimise peptide and protein identification. Differential gene and protein expression was assessed using linear models correcting for age, gender, age x gender interaction, and unwanted variation. FDR