3′-tRF-Cys(GCA) overexpression in HEK-293 cells alters the global expression profile and modulates cellular processes and pathways

tRNA fragments (tRFs) are small non-coding RNAs generated through specific cleavage of tRNAs and involved in various biological processes. Among the different types of tRFs, the 3′-tRFs have attracted scientific interest due to their regulatory role in gene expression. In this study, we investigated the role of 3′-tRF-Cys(GCA), a tRF deriving from cleavage in the T-loop of tRNA(CysGCA), in the regulation of gene expression in HEK-293 cells. Previous studies have shown that 3′-tRF-Cys(GCA) is incorporated into the RISC complex and interacts with Argonaute proteins, suggesting its involvement in the regulation of gene expression. However, the general role and effect of the deregulation of 3′-tRF-Cys(GCA) levels in human cells have not been investigated so far. To fill this gap, we stably overexpressed 3′-tRF-Cys(GCA) in HEK-293 cells and performed transcriptomic and proteomic analyses. Moreover, we validated the interaction of this tRF with putative targets, the levels of which were found to be affected by 3′-tRF-Cys(GCA) overexpression. Lastly, we investigated the implication of 3′-tRF-Cys(GCA) in various pathways using extensive bioinformatics analysis. Our results indicate that 3′-tRF-Cys(GCA) overexpression led to changes in the global gene expression profile of HEK-293 cells and that multiple cellular pathways were affected by the deregulation of the levels of this tRF. Additionally, we demonstrated that 3′-tRF-Cys(GCA) directly interacts with thymopoietin (TMPO) transcript variant 1 (also known as LAP2α), leading to modulation of its levels. In conclusion, our findings suggest that 3′-tRF-Cys(GCA) plays a significant role in gene expression regulation and highlight the importance of this tRF in cellular processes. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s10142-023-01272-0.