Unraveling the Role of tRNA-Derived Fragments in Post-Transcriptional Gene Regulation

Authors: Merve Nisa Çakır, İlke Aksoy, & Abu Musa Md Talimur Reza

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In the classical central dogma of molecular biology, the process of gene expression is a straightforward path: DNA is transcribed into RNA, which is then translated into proteins. However, recent scientific breakthroughs have revealed that this linear model doesn't fully explain the complexity of gene regulation. One of the most exciting discoveries in recent years is the role of non-coding RNAs, particularly tRNA-derived fragments (tRFs) and tRNA halves (tiRNAs), in the post-transcriptional control of gene expression.

These small RNA molecules, once dismissed as degradation products, are now recognized as crucial regulatory players with diverse roles in health, development, and disease.

Beyond Translation: The Hidden Life of tRNAs

Transfer RNAs (tRNAs) are best known for their central role in translation—delivering amino acids to the ribosome during protein synthesis. But they are far more than just passive adaptors. In human cells, tRNAs (~72 nucleotides in length) are encoded by hundreds of genes and undergo extensive post-transcriptional processing and chemical modifications.

Under normal or stress conditions, mature and precursor tRNAs can be precisely cleaved into smaller fragments ranging from 18 to 50 nucleotides. These tRNA-derived small RNAs (collectively termed tsRNAs) are biologically active molecules that regulate mRNA stability, translation, and even transcription.

Biogenesis of tRNA-Derived Fragments

1. tRNA Halves (tiRNAs)

tiRNAs are generated by cleavage in the anticodon loop of mature tRNAs and are typically produced during cellular stress. This cleavage is primarily catalyzed by angiogenin, a stress-responsive endonuclease. tiRNAs are classified into:

• 5′-tiRNAs: from the 5′ portion of the tRNA

• 3′-tiRNAs: from the 3′ portion

These fragments can suppress global translation and promote stress granule formation, acting as part of the cell’s emergency response system.

2. tRNA-Derived Fragments (tRFs)

tRFs are shorter than tiRNAs and arise from precise cleavage at different tRNA regions:

• tRF-5: from the 5′ end of mature tRNAs (often Dicer-dependent)

• tRF-3: from the 3′ end (TψC loop) of mature tRNAs (angiogenin-/Dicer-mediated)

• tRF-1: from the 3′ trailer sequence of precursor tRNAs (processed by RNase Z)

• Internal tRFs (i-tRFs): from internal regions of mature tRNAs; their biogenesis is less well understood

These categories are functionally diverse, and ongoing research is revealing their involvement in RNA silencing, translation regulation, and beyond.

How Do tRFs and tiRNAs Regulate Gene Expression?

1. RNA Silencing via Argonaute Proteins

   Much like microRNAs, several tRFs—especially tRF-3s—associate with Argonaute (Ago) proteins to guide gene silencing. tRFs can bind complementary mRNA sequences to inhibit translation or trigger mRNA degradation. Interestingly, they show distinct preferences for specific Ago isoforms, often binding more effectively to Ago3 and Ago4 than to Ago1 or Ago2.

2. Positive Translation Regulation

   Not all tRFs are repressors. A remarkable example is a 3′-tsRNA derived from tRNA^Leu-CAG, which binds to the structured 5′ UTR of rps28 mRNA, promoting its translation and contributing to ribosome biogenesis.

3. Modulating Histone mRNA Stability

   Histone mRNAs are unique as they lack poly-A tails and are regulated differently from typical mRNAs. Some tRF-3s bind near the stem-loop structures of histone mRNAs, potentially interfering with SLBP (stem-loop binding protein) and altering mRNA stability.

4. Nuclear Functions and Alternative Splicing

   Although traditionally thought to function in the cytoplasm, Argonaute proteins and tRF-5s are also found in the nucleus. This suggests that tRFs may influence transcriptional repression or alternative splicing—a fascinating area for future research.

Piwi Protein Interactions: A piRNA-Like Role?

Beyond Argonautes, certain longer tRFs (26–35 nt) bind Piwi proteins, suggesting a piRNA-like function. This interaction is observed across species—from Drosophila to human cells. In humans, Hiwi2, a Piwi protein, has been shown to bind 5′-tsRNAs and co-sediment with polysomes, hinting at a role in translation regulation in cancer cells.

Implications and Future Directions

The growing recognition of tRFs and tiRNAs as functional regulators reshapes our understanding of gene expression. These small RNAs are involved in: stress response pathways, mRNA stability and translation, cellular differentiation and development, and cancer progression and metastasis. Despite this progress, many questions remain:

• What are the precise molecular mechanisms of tRF action?

• How are these fragments selectively produced and regulated?

• Can they serve as reliable biomarkers or therapeutic targets?

Conclusion

Once overlooked as byproducts of RNA degradation, tRNA-derived fragments are now emerging as essential players in gene regulation. Their versatility and involvement in both gene repression and activation suggest a broader regulatory scope than previously imagined. As research continues, tRFs and tiRNAs may soon join the ranks of microRNAs and lncRNAs as central regulators in cellular biology and disease.