Type

Journal Article

Authors

Richard Murphy
Padraig Doolan
Karina Horgan
Martin Clynes
Niall Barron
Laura Breen
Paula Meleady
Michael Henry
Colin Clarke
Fiona O'Neill
and 3 others

Subjects

Biochemistry

Topics
mrna expression cell differentiation gene expression cell adhesion hierarchical cluster analysis cell line cell models cell lines

Parallel mRNA, proteomics and miRNA expression analysis in cell line models of the intestine. (2017)

Abstract To identify miRNA-regulated proteins differentially expressed between Caco2 and HT-29: two principal cell line models of the intestine. Exponentially growing Caco-2 and HT-29 cells were harvested and prepared for mRNA, miRNA and proteomic profiling. mRNA microarray profiling analysis was carried out using the Affymetrix GeneChip Human Gene 1.0 ST array. miRNA microarray profiling analysis was carried out using the Affymetrix Genechip miRNA 3.0 array. Quantitative Label-free LC-MS/MS proteomic analysis was performed using a Dionex Ultimate 3000 RSLCnano system coupled to a hybrid linear ion trap/Orbitrap mass spectrometer. Peptide identities were validated in Proteome Discoverer 2.1 and were subsequently imported into Progenesis QI software for further analysis. Hierarchical cluster analysis for all three parallel datasets (miRNA, proteomics, mRNA) was conducted in the R software environment using the Euclidean distance measure and Ward's clustering algorithm. The prediction of miRNA and oppositely correlated protein/mRNA interactions was performed using TargetScan 6.1. GO biological process, molecular function and cellular component enrichment analysis was carried out for the DE miRNA, protein and mRNA lists Differential expression (DE) profiling comparing the intestinal cell lines HT-29 and Caco-2 identified 1795 Genes, 168 Proteins and 160 miRNAs as DE between the two cell lines. At the gene level, 1084 genes were upregulated and 711 were downregulated in the Caco-2 cell line relative to the HT-29 cell line. At the protein level, 57 proteins were found to be upregulated and 111 downregulated in the Caco-2 cell line relative to the HT-29 cell line. Finally, at the miRNAs level, 104 were upregulated and 56 downregulated in the Caco-2 cell line relative to the HT-29 cell line. Gene ontology (GO) analysis of the DE mRNA identified cell adhesion, migration and ECM organization, cellular lipid and cholesterol metabolic processes, small molecule transport and a range of responses to external stimuli, while similar analysis of the DE protein list identified gene expression/transcription, epigenetic mechanisms, DNA replication, differentiation and translation ontology categories. The DE protein and gene lists were found to share 15 biological processes including for example epithelial cell differentiation [ This first study providing "tri-omics" analysis of the principal intestinal cell line models Caco-2 and HT-29 has identified 34 proteins potentially undergoing miRNA translational repression.
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Full list of authors on original publication

Richard Murphy, Padraig Doolan, Karina Horgan, Martin Clynes, Niall Barron, Laura Breen, Paula Meleady, Michael Henry, Colin Clarke, Fiona O'Neill and 3 others

Experts in our system

1
Padraig Doolan
Dublin City University
Total Publications: 37
 
2
Martin Clynes
Dublin City University
Total Publications: 209
 
3
Niall Barron
Dublin City University
Total Publications: 44
 
4
Laura Breen
Dublin City University
Total Publications: 11
 
5
Paula Meleady
Dublin City University
Total Publications: 95
 
6
Michael Henry
Dublin City University
Total Publications: 78
 
7
Colin Clarke
Dublin City University
Total Publications: 26
 
8
Fiona O'Neill
Dublin City University
Total Publications: 4