Exploring ribosome composition and newly synthesized proteins through proteomics and potential biomedical applications

Abstract

Protein synthesis is the outcome of tightly regulated gene expression which is responsive to a variety of conditions. Efforts are ongoing to monitor individual stages of protein synthesis to ensure maximum efficiency and accuracy. Due to post-transcriptional regulation mechanisms, the correlation between translatome and proteome is higher than between transcriptome and proteome. However, the most accurate approach to assess the key modulators and final protein expression is directly by using proteomics. Areas covered: This review covers various proteomic strategies that were used to better understand post-transcriptional regulation, specifically during and early after translation. The methods that identify both regulatory proteins associated with translational components and newly synthesized proteins are discussed. Expert commentary: Emerging proteomic approaches make it possible to monitor protein dynamics in cells, tissues and whole animals. The ability to detect alteration in protein abundance soon after their synthesis enables earlier recognition of disease causing factors and candidates to prevent/rectify disease phenotype.

Keywords: AHA; Post-transcriptional regulation; dynamic proteomics; fluorescent imaging of synthesized proteins; mass spectrometry; newly synthesized proteins; non-canonical amino-acid labeling; ribosomes; translation.

Figure 1.

Nuclear encoded protein expression. Transcription – information from DNA template in nucleus is transcribed to RNA (pre-mRNA; RNA transcript) that is further processed into functional messenger mRNA by capping, splicing (introns removed and exons ligated) and polyadenylation. Mature mRNA containing codons is exported into cytoplasm via nuclear pore complex. Various RNA-binding proteins, RNA-associated proteins (spliceosome, small nuclear snRNPs), splicing and transcription factors and accessory proteins regulate this step. In cytoplasm, mRNA becomes associated with multiple ribosomes forming polysomes. The amino acids (aa_s) present in cytoplasm are activated, attached to proper transfer tRNAs containing anti-codons in the presence of aminoacyl-tRNA-synthetase (tRNAs present in cytoplasm are “charged”) and transported to mRNA on ribosome. At the beginning of protein synthesis, the ribosome is positioned over a start codon of mRNA and polypeptide synthesis takes place. The amino acids are transferred to growing peptide chain as mRNA moves through the ribosome and empty (“uncharged”) tRNAs return to cytoplasm. After polypeptide is completed, it is released to cytoplasm, subjected to post-translational processing into active protein (folding, splicing, post-translationally modificated…) and ribosome subunits separate. During this post-transcriptional steps, many regulatory RNAs, RNA-binding and associated proteins are present as well as various translation factors and chaperones. The methods and approaches for identification/quantitation mRNA/proteins are described in detail throughout the text.