Translatomics for Isodecoders, pre-tRNAs and plant seeds
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,
- Alarcon, R. et al. design the multiplexed long read tRNAseq method
- Verstraten, R. et al. develop a long read approach for sequencing pre-tRNAs
- Bai B. et al. present the translational landscape at sub-codon resolution across Arabidopsis seed germination
ADAM-tRNA-seq: an optimized approach for demultiplexing and enhanced hierarchal mapping in direct tRNA sequencing
Nucleic Acid Research, 2026
Alarcon, R., Köster, D., Behrmann, S. and Ignatova, Z.
The study introduces ADAM-tRNA-seq, a framework designed to refine direct RNA sequencing (DRS) via the Nanopore platform for tRNA-seq. While DRS theoretically allows for the simultaneous detection of tRNA abundance, aminoacylation status, and base modifications, it has historically been plagued by low accuracy due to the high sequence similarity between tRNA isodecoders and the lack of robust demultiplexing tools for small, highly structured RNAs.
ADAM-tRNA-seq overcomes these hurdles through two primary innovations. The first one is integrated barcoding: by embedding barcodes directly into the RNA adapters, the method achieves near-perfect (up to 99% precision) demultiplexing, essential for scaling experiments.
Cellular tRNAs exhibit high similarity between isodecoders and also they are highly modified which creates basecalling errors. Therefore, the second innovation handles that with hierarchical mapping workflow. For instance, if comparison of the abundances of isodecoders is of interest, then only reads that can be unambiguously identified as a specific isodecoder should be kept. If distribution of isoacceptors is the focus of the analysis, then reads that multimap to different isodecoders within the same isoacceptor family can be kept and pooled. Such an approach significantly reduces mapping biases that previously obscured the true composition of the tRNA pool. The authors optimized and tested their approach on vitro-transcribed tRNAs and human HEK293 cells.
By providing a high-resolution full-length snapshot of the tRNA pool alongside modification patterns, ADAM-tRNA-seq enables a deeper understanding of how tRNA dynamics regulate protein synthesis rates, ultimately linking tRNAome dysregulation to human pathologies.
Defining expansions and perturbations to the RNA polymerase III transcriptome and epitranscriptome by modified direct RNA nanopore sequencing
Nature Communications, 2026
Verstraten, R., Cetraro, P., Fitzpatrick, A.H. et al.
The study introduces DRAP3R, a sequencing approach designed to improve the characterization of RNA polymerase III (Pol III) transcripts, including tRNAs, which are difficult to study using conventional RNA-sequencing techniques. The authors developed DRAP3R, a nanopore-based direct RNA sequencing strategy that enables simultaneous detection of Pol III transcripts and their chemical modifications. One of the key protocol steps includes depletion of non-poly(U) RNAs prior to ligation with a custom poly(U)-targeting adapter thus enabling the capture of pre-mature poly(U)-tailed RNAs, which arise from Pol III transcription termination. This method captures full-length RNA molecules without the need for reverse transcription or amplification, reducing biases that commonly affect tRNA-seq datasets. By sequencing native RNA molecules directly, the approach preserves information about RNA modifications, which are crucial regulators of tRNA stability, structure, and function.
Using this technology, the researchers generated high-resolution datasets of Pol III-derived RNAs, including multiple tRNA species. Their analysis validated previously uncharacterized RNAs that were inferred from the ChIP-Seq binding profiles of select Pol III subunits. Also, with this approach it was revealed that pre-tRNAs carry pseudouridine modifications. It is known that HSV-1 infection leads to changes in pre-tRNA and mature tRNA levels however it is not yet known whether pseudouridine installation is regulated. To address this, researchers applied DRAP3R to RNA collected from two biological replicates each of HSV-1 infected ARPE-19s collected at 6 h post infection (hpi) and 12 hpi. It revealed an extensive remodelling of both the Pol III transcriptome and epitranscriptome including a significant upregulation in expression of ~50 pre-tRNAs and significant downregulation of most Pol II transcribed genes at both 6 and 12 hpi.
Overall, this work highlights the potential of nanopore-based tRNA-seq approaches to uncover previously hidden layers of RNA regulation, offering new opportunities to study tRNA function, RNA modification dynamics, and the broader role of Pol III transcripts in gene expression.
Learn more about EIRNABio’s tRNA-seq profiling services here.
Translational landscape during seed germination revealed by ribosome profiling
The Plant Journal, 2026
Bai B., Qi R., Song W., Nijveen H., Bentsink L.
Seed germination is crucial for agricultural reproduction and better understanding of the biology of this process can help to improve plant growth and increase yield. To investigate the mechanism of translational control, the scientists employed ribosome profiling on mRNAs of five distinct physiological stages during Arabidopsis thaliana seed germination including dry seeds, seeds at early imbibition, seeds before testa rupture, seeds at 80% of endosperm rupture and 80% of seedling greening.
In Arabidopsis, mRNAs stored in dry seeds are predominantly associated with monosomes that are not actively involved in translation. Interestingly, in these translationally inactive dry seeds, researchers found a unique ribosome association landscape with a higher ribosome association at the 5′ and 3′ UTR, compared with physiological stages that show active translation. In addition, they discovered an adenine-enriched sequence motif that is linked to a start codon-specific stalling of ribosomes in dry seeds.
In the next germination stages, it was shown that codons encoding glycine, aspartate, tyrosine, and proline are the most frequent ribosome pausing sites which is consistent with previous observations showing that proline, glycine, and aspartate are the most conserved amino acids associated with ribosome stalling in yeast, fruit fly, zebrafish, mouse, and human. Moreover, non-coding RNAs were found to be associated with ribosomes and were identified as being actually translated as it was further endorsed by proteomic data – in total, 50 out of 426 lncRNA‐encoded peptides were identified in the seed total proteome during germination. Also, non-canonical translons such as uORFs have been found that may play a role in translational regulation of early seed germination. For instance, the translation of the uORF of ABH (AT5G36210) in dry seeds represses the translation of the ABH mORF, whereas upon imbibition this repression is attenuated. The ABH domain can serve as a core component in the perception of various compounds promoting seed germination.
Learn more about EIRNABio’s ribosome profiling services here.
