Abstract
Transfer RNA-derived fragments (tRFs) are emerging small noncoding RNAs that, although commonly altered in cancer, have poorly defined roles in tumorigenesis1. Here we show that pseudouridylation (Ψ) of a stem cell-enriched tRF subtype2, mini tRFs containing a 5′ terminal oligoguanine (mTOG), selectively inhibits aberrant protein synthesis programmes, thereby promoting engraftment and differentiation of haematopoietic stem and progenitor cells (HSPCs) in patients with myelodysplastic syndrome (MDS). Building on evidence that mTOG-Ψ targets polyadenylate-binding protein cytoplasmic 1 (PABPC1), we employed isotope exchange proteomics to reveal critical interactions between mTOG and functional RNA-recognition motif (RRM) domains of PABPC1. Mechanistically, this hinders the recruitment of translational co-activator PABPC1-interacting protein 1 (PAIP1)3 and strongly represses the translation of transcripts sharing pyrimidine-enriched sequences (PES) at the 5′ untranslated region (UTR), including 5′ terminal oligopyrimidine tracts (TOP) that encode protein machinery components and are frequently altered in cancer4. Significantly, mTOG dysregulation leads to aberrantly increased translation of 5′ PES messenger RNA (mRNA) in malignant MDS-HSPCs and is clinically associated with leukaemic transformation and reduced patient survival. These findings define a critical role for tRFs and Ψ in difficult-to-treat subsets of MDS characterized by high risk of progression to acute myeloid leukaemia (AML).
Highlights
A prominent Ψ ‘writer’ responsible for stress-inducible Ψ on different RNAs is the multi-substrate synthase PUS7
We performed an electrophoretic mobility shift assay to determine the binding dynamics. This experiment unambiguously demonstrated that mini tRFs containing a 5′ terminal oligoguanine (mTOG) directly engaged polyadenylate-binding protein cytoplasmic 1 (PABPC1) in a Ψ-dependent manner, as revealed by a lower dissociation constant compared with unmodified mTOG or Ψ-modified scramble (SCR-Ψ) oligos (Fig. 1a and Extended Data Fig. 1a)
Given that translation of 5′ pyrimidine-enriched sequences (PES)-containing messenger RNA (mRNA) is highly responsive to the assembly and function of the cap-binding translation initiation complex, we evaluated the direct contribution of mTOG-Ψ to the translation of selected transcripts identified through ribosome profiling in PUS7-KO cells
Summary
Consistent with our translation analysis, the protein levels of RPL29, RPL23 and EIF6 were drastically increased in PUS7-KO hESCs and could be restored only following transfection of mTOG-Ψ (Fig. 2e) These effects were independent of changes in mRNA transcription and stability, and were uncoupled from differences in the precursor and mature tRNA-Tyr(GUA), a well-established PUS7 substrate in eukaryotic cells[6] (Fig. 2e and Extended Data Fig. 2g,h). Depletion of either PABPC1 or PAIP1 selectively rescued increased translation of these mRNAs with no additional mTOG-Ψ-dependent translation repression observed (Fig. 2g and Extended Data Fig. 3a). We found that mTOG-Ψ treatment further repressed protein synthesis in LARP1-depleted PUS7-KO cells (Extended Data Fig. 3c) This indicates that mTOG and LARP1 regulate PABPC1 and translation through a distinct mechanism, consistent with findings that mTOG-Ψ does not affect LARP1 binding to PABPC1 (Fig. 1d). Consistent with previous observations that increased protein synthesis downstream of eIF4F is associated with the activation a cDNA Sequencing and mapping b log2(PUS7-KO TE) Cumulative proportion
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
