Identifying mRNA subsets in messenger ribonucleoprotein complexes by using cDNA arrays

TLDR

Genomic array technologies enable profiling of global gene expression changes, but distinguishing transcriptional from posttranscriptional regulation and detecting cell‑type specific variations within heterogeneous tissues remains challenging. The study aims to use cDNA arrays to identify cell‑type specific mRNA subsets within endogenous messenger ribonucleoprotein complexes (mRNPs) by recovering associated proteins. By immunoprecipitating mRNA‑binding proteins HuB, eIF‑4E, and PABP from P19 embryonal carcinoma stem cells and profiling the co‑precipitated mRNAs with cDNA arrays, the authors map mRNP‑associated transcript clusters. The mRNP‑associated mRNA profiles were distinct from total cellular RNA, altered dramatically upon retinoic‑acid‑induced neuronal differentiation, and suggest that dynamic clustering of related mRNAs regulates posttranscriptional events, establishing ribonomics as a useful functional genomics paradigm.

Abstract

Genomic array technologies provide a means for profiling global changes in gene expression under a variety of conditions. However, it has been difficult to assess whether transcriptional or posttranscriptional regulation is responsible for these changes. Additionally, fluctuations in gene expression in a single cell type within a complex tissue like a tumor may be masked by overlapping profiles of all cell types in the population. In this paper, we describe the use of cDNA arrays to identify subsets of mRNAs contained in endogenous messenger ribonucleoprotein complexes (mRNPs) that are cell type specific. We identified mRNA subsets from P19 embryonal carcinoma stem cells by using mRNA-binding proteins HuB, eIF-4E, and PABP that are known to play a role in translation. The mRNA profiles associated with each of these mRNPs were unique and represented gene clusters that differed from total cellular RNA. Additionally, the composition of mRNAs detected in HuB–mRNP complexes changed dramatically after induction of neuronal differentiation with retinoic acid. We suggest that the association of structurally related mRNAs into mRNP complexes is dynamic and may help regulate posttranscriptional events such as mRNA turnover and translation. Recovering proteins specifically associated with mRNP complexes to identify and profile endogenously clustered mRNAs should provide insight into structural and functional relationships among gene transcripts and/or their protein products. We have termed this approach to functional genomics ribonomic s and suggest that it will provide a useful paradigm for organizing genomic information in a biologically relevant manner.