Leveraging the complementary nature of RNA‐Seq and shotgun proteomics data

Abstract

RNA sequencing (RNA-Seq) and mass spectrometry-based shotgun proteomics are powerful high-throughput technologies for identifying and quantifying RNA transcripts and proteins respectively. With the increasing affordability of these technologies, many projects have started to apply both to the same samples to achieve a more comprehensive understanding of biological systems. A major analytical challenge for such integrative projects is how to effectively leverage the complementary nature of RNA-Seq and shotgun proteomics data. RNA-Seq provides comprehensive information on mRNA abundance, alternative splicing, nucleotide variation and structure alteration. Sample-specific protein databases derived from RNA-Seq data can better approximate the real protein pools in cell and tissue samples and thus improve protein identification. Meanwhile, proteomics data provides essential confirmation of the validity and functional relevance of novel findings from RNA-Seq data. At the quantitative level, mRNA and protein levels are only modestly correlated, suggesting strong involvement of post-transcriptional regulation in controlling gene expression. Here we review recent studies at the interface of RNA-Seq and proteomics data. We discuss goals, accomplishments and challenges in RNA-Seq-based proteogenomics. We also examine the current status and future potential of parallel transcriptome and proteome quantification in revealing post-transcriptional regulatory mechanisms.

Slowed decay of mRNAs enhances platelet specific translation

“…61,[69][70][71] We next sought to better understand this discordance using our ribosome profiling data and determine the extent to which it may reflect biological phenomena such as uptake/sequestration of serum proteins, 72 or other unusual features of platelet gene expression, 13 vs technical artifacts of the different methods used to measure nucleic acids and proteins. 73,74 High-confidence genes identified by all 3 methods (RNA-Seq, ribosome profiling, and mass spectrometry), which were both synthesized and accumulated in platelets, were significantly enriched for cytoplasmic processes such as "intracellular transport" and "gene expression" (q value , 0.05) ( Figure 2C) and depleted for nuclear functions, consistent with the lack of nuclei in platelets (supplemental Tables 1-3). In contrast, genes that were detected by RNA-Seq and ribosome profiling, but not by mass spectrometry, were enriched in nuclear components and functions such as transcription and chromatin organization (supplemental Figure 2A; supplemental Tables 4-6), perhaps suggesting rapid degradation of unneeded nuclear proteins in platelets.…”

Integrative Omics Analysis Reveals Post-Transcriptionally Enhanced Protective Host Response in Colorectal Cancers with Microsatellite Instability

“…A similar correlation coefficient value has been observed by many previous comparative transcriptomics and proteomics studies independent of the proteomics approach (labeling or label-free) and quantification method (spectral count or intensity-based). 62,63 Therefore, the divergence between mRNA and protein is more likely to be driven by post-transcriptional regulation rather than an artifact introduced by the protein quantification method. A simple comparison between differentially expressed mRNAs and proteins (|log2FC| > 1 and adjusted p -value <0.05) identified only 26 common upregulated genes and 19 downregulated genes (Figure 4A).…”

Linking transcriptomics and proteomics in spermatogenesis

“…Their transcriptomes and proteomes can be thoroughly examined without requiring the use of nucleic or protein sequence databases from phylogenetically distant species. Finally, regardless of the model of interest, the combination of RNA-seq and mass spectrometry into a PIT study offers a straightforward method of investigating the correlation of transcriptomes and proteomes, because protein profiles can be directly compared to transcript profiles on which protein identifications are also performed (for review, see Wang et al (2014)). …”