September 3rd
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, Zhulyn et al. (2023) show a distinct characteristic of the injury response necessary for limb regeneration in the axolotl. Munro et al. (2023) provide sample lysis conditions that are optimized for enabling the precise and consistent application of techniques for studying translational control in primary mouse tissue. Lastly, Li and Chen (2023) use genome-wide ribosome profiling data and machine learning models to envisage SNP function by predicting ribosome collisions during mRNA translation.
Evolutionarily divergent mTOR remodels translatome for tissue regeneration
Nature, 2023
Zhulyn, O., Rosenblatt, H.D., Shokat, L., Dai, S., Kuzuoglu-Öztürk, D., Zhang, Z., Ruggero, D., Shokat, K.M. and Barna, M.
Why some species, like the axolotl (Ambystoma mexicanum), can regenerate tissues but mammals cannot largely remains to be seen. De novo transcription and processing of axolotl transcripts may not be adequate to investigate rapid responses, including wound healing following limb amputation, which take place within hours of injury. A deeper comprehension of the molecular mechanisms controlling gene expression in quick wound healing is needed. Here, the authors show that a distinct characteristic of the injury response necessary for limb regeneration in the axolotl is the rapid activation of protein synthesis. They employed polysome sequencing to report hundreds of transcripts, including antioxidants and ribosome components, which are specifically triggered at the translational level in response to injury from pre-existing messenger RNAs. In contrast, when a mouse has a digit amputated without regeneration, protein synthesis is not triggered.
They propose the mTORC1 pathway as a crucial upstream signal that regulates translational regulation and tissue regeneration in the axolotl and find that urodele amphibians have particular expansions in the mTOR protein sequence. These modifications result in a hypersensitive kinase that enables axolotls to retain this pathway in a highly labile state prepared for prompt activation. Together, these findings fill in a critical gap in the knowledge of the regenerative potential of vertebrates.
Optimisation of sample preparation from primary mouse tissue to maintain RNA integrity for methods examining translational control
Cancers, 2023
Munro, J., Gillen, S.L., Mitchell, L., Laing, S., Karim, S.A., Rink, C.J., Waldron, J.A. and Bushell, M.
The amount of protein that an mRNA produces depends on how quickly it is decoded by the ribosome, which can differ significantly between mRNAs. Methods for measuring the translation rates of particular mRNAs cannot be denaturing-based, since native mRNA-ribosome interactions must be preserved. Key techniques for determining translation rates taking place on certain mRNAs include polysome profiling and ribosome profiling. Although these methods are now often utilized in cell lines, primary tissues are frequently found to have higher levels of ribonuclease (RNase) expression. Here, the authors provide sample lysis conditions that are optimized for non-denaturing circumstances, enabling the precise and consistent application of techniques for studying translational control in primary mouse tissue.
The authors tested the ability of different RNase inhibitors to maintain the integrity of RNA from a variety of tissues that express endogenous RNase at various amounts. Heparin increased the integrity of the liver, lung, and colon but was insufficient for the small intestine, while SUPERaseIn very minimally improved integrity across all tissue types. The most consistent increases in RNA integrity across the tissues were produced by RiboLock and the murine RNase inhibitor. None of the RNase inhibitors tested were able to effectively prevent RNA degradation in the pancreas owing to the high levels of endogenous RNases present, limiting the application of translatomics approaches in in this tissue. The authors further emphasize the significance of RNA integrity for the accurate calculation of the global translation status of the cell as assessed by polysome gradients as failure to check RNA integrity across all collected polysome fractions can lead to a misinterpretation of results. Moreover, polysome gradient profiles demonstrated how different tissues have very different levels of ribosome occupancy, highlighting the importance of being able to accurately study translation in a wide number of tissues.
Predicting functional consequences of SNPs on mRNA translation via machine learning
Nucleic Acids Research, 2023
Li, Z. and Chen, L.
Prioritizing disease-causing single nucleotide polymorphisms (SNPs) from genome-wide association studies (GWAS) has not yet taken the functional impact of SNPs on translation into account. Finding the causative variations of common diseases remains difficult. Therefore, to effectively utilize the power of GWAS in disease diagnosis and treatment discovery, it is necessary to comprehend the functional and clinical effects of genetic variations. Here, the authors use genome-wide ribosome profiling data and machine learning models to project SNP function by predicting ribosome collisions during mRNA translation. RibOc-SNPs (Ribosome-Occupancy-SNPs) are SNPs that significantly alter ribosome occupancy. They discovered that disease-related SNPs frequently result in noticeable alterations in ribosome occupancy, indicating that translational regulation is a critical stage in pathogenesis.
Ribosome occupancy is most significantly affected by nucleotide conversions enriched in RibOc-SNPs such as ‘G→T,’ ‘T→G,’ and ‘C→A,’ however ‘A→G’ (or ‘A→I’ RNA editing) and ‘G→ A” are less deterministic. The conversion ‘Glu→stop (codon)’ exhibits the greatest enrichment in RibOc-SNPs when compared to other amino acid transformations. It’s interesting to note that stop codons with a decreased risk of collision are under selection pressure. The 5′-coding sequence areas are richer in ribOc-SNPs, suggesting that these are hotspots for the control of translation initiation. Surprisingly, 22.1% of the RibOc-SNPs cause opposing changes in ribosome occupancy on alternative transcript isoforms. This finding raises the possibility that SNPs can accentuate the disparities between splicing isoforms by oppositely affecting their translation efficiency.