September 17th

Recent Publications Harnessing the Power of Translatomics.

Every week we provide a digest of a small number of recent interesting papers in the field of translatomics.

In this week’s Sunday papers, Cuevas et al. (2023) discuss their tool, BamQuery, designed to analyse MHC-I-associated peptides with lengths between 8 to 11 amino acids. Bowazolo & Morse (2023) investigate how light-dark cycles have the ability to synchronise the translation in Fugacium kawagutii, finally, Dong et al. characterised 4EBP1 and 4EBP2 proteins in Arabidopsis.

BamQuery: a proteogenomic tool to explore the immunopeptidome and prioritize actionable tumor antigens

Genome Biology, 2023

Ruiz Cuevas, M. V., Hardy, M.-P., Larouche, J.-D., Apavaloaei, A., Kina, E., Vincent, K., Gendron, P., Laverdure, J.-P., Durette, C., Thibault, P., Lemieux, S., Perreault, C., & Ehx, G.

The immunopeptidome captures MHC-I-associated peptides (MAPs) reflecting intracellular mRNA translation and degradation. This data aids in identifying targetable tumour antigens (TAs), including neoantigens from mutation-driven neoplasms. Around 5-10% of MAPs arise from non-canonical genomic regions like introns, ncRNAs, endogenous retroelements, and out-of-frame exonic translation. These non-coding and mutation-based MAPs yield tumour-specific antigens. BamQuery, introduced here, assigns comprehensive RNA expression to MAPs from diverse sources using RNA-sequencing data. It uncovers cryptic and mutated TAs from multiple highly expressed genomic regions in normal tissues, aiding in TA immunogenicity prediction and de novo TA identification. BamQuery is a tool designed to analyse MAPs with lengths between 8 to 11 amino acids.

To investigate the relationship between the transcriptome and immunopeptidome, the study capitalized on data from three diffuse large B cell lymphoma cell lines, equipped with matched RNA-seq, Ribo-seq, and immunopeptidomic data. Transcripts yielding MAPs were abundant compared to non-source transcripts, affirming RNA expression’s predictive role in MAP generation likelihood. Leveraging ribo-seq data, aligned reads formed bam files for BamQuery analysis. Correlating RNA-seq and Ribo-seq quantifications for each identified MAP exhibited strong concordance, validating RNA expression as a reliable indicator of MAP translation likelihood.

BamQuery ensures consistent validation of tumor antigens (TAs) across different research settings, uncovering a notable fraction of the immunopeptidome originating from non-coding regions, highlighting the importance of the “dark genome.” BamQuery’s contribution lies in standardizing proteogenomic analyses in MHC-I immunopeptidomics, allowing prioritization of TAs lacking RNA expression in normal tissues, making them safer for clinical trial testing.

Ribosome profiling in the Symbiodiniacean dinoflagellate Fugacium kawagutii shows coordinated protein synthesis of enzymes in different pathways at different times of day

Molecular Microbiology., 2023

Bowazolo, C. and Morse, D.

Dinoflagellates exhibit responses to diurnal variations in light and darkness through alterations in cellular metabolism. However, the precise mechanisms underlying these responses remain elusive, as previous attempts to identify rhythmically expressed transcripts via RNA Seq experiments was unsuccessful, suggesting that neither fluctuations in light intensity nor circadian clock control depend on transcriptional mechanisms altering mRNA levels.

In this study, ribosome profiling was conducted at 2-hour intervals throughout a daily light-dark cycle to evaluate potential fluctuations in protein synthesis rates in Fugacium (formerly known as Symbiodinium) kawagutii. The analysis revealed that a total of 2,923 coding sequences of F. kawagutii exhibited significant variations in the number of ribosome-protected fragments (RPF) over the 24-hour cycle. Notably, the study identified concurrent regulation of numerous transcripts encoding proteins involved in spliceosome or lysosome biogenesis. Each of these regulated transcripts exhibited peak translation activity at the onset of the dark phase.

Conversely, transcripts associated with nitrate metabolism and ribosomal proteins were preferentially translated around the onset of the day phase or the conclusion of the night phase, respectively. Additionally, some sequences involved in DNA synthesis exhibited a preference for translation at the end of the day.

In conclusion, this study suggests that light-dark cycles have the ability to synchronise the translation of specific transcripts encoding proteins engaged in diverse cellular processes. These changes in translation may aid cells in adapting and modifying their metabolism in response to the time of day.

Functional analogs of mammalian 4E-BPs reveal a role for TOR in global plant translation

Cell Reports, 2023

Dong, Y., Srour, O., Lukhovitskaya, N., Makarian, J., Baumberger, N., Galzitskaya, O., Elser, D., Schepetilnikov, M., & Ryabova, L. A.

The mTOR protein regulates protein synthesis by deactivating eIF4E-binding proteins (m4E-BPs) in response to nutrient and energy levels. Until recently, it was believed that 4E-BPs were unique to animals, and how TOR controls cap-dependent translation initiation in plants was not understood. The aim of this study was then to investigate how the target of rapamycin (TOR) regulates cap-dependent translation initiation in plants using techniques which included polysome profiling and Translating ribosome affinity immunopurification (TRAP)-seq.

Here, the authors identified short unstructured proteins in Arabidopsis (4EBP1 and 4EBP2) that share similarities with eIF4E-binding proteins (eIF4E-BPs). These proteins were found to contain the eIF4E-binding motif and TOR phosphorylation sites. Unphosphorylated 4EBPs were found to have the ability to inhibit the formation of the cap-binding complex, which is essential for translation initiation.

The authors discovered that 4EBPs could selectively suppress the translation of specific mRNAs. For example, 4EBP2 predominantly inhibited the translation of core cell-cycle regulators, while 4EBP1 interfered with chlorophyll biosynthesis. Overexpression of 4EBP2 had a notable effect on early seedling development, which was overcome by the induction of Glc/Suc-TOR signaling. This suggests that TOR regulates cap-dependent translation initiation through the inactivation of these atypical 4EBPs in plants.

In summary, the authors identified and characterized 4EBP1 and 4EBP2 proteins in Arabidopsis, elucidated their role in regulating translation initiation through their interaction with eIF4Es and their phosphorylation by TOR, and demonstrated their specific effects on the translation of certain mRNAs. These findings contribute to a better understanding of how TOR regulates protein synthesis in plants.

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