bioRxiv Science⌕ Search
Home/Archive/bioRxiv

bioRxiv · 10.64898/2026.05.28.728624

m1A58 governs two steps of human initiator-methionine tRNA metabolism: maturation and XRN2-mediated decay

Otsubo, K.·Nagura, M.·Terauchi, M.·Noguchi, H.·Kanemaki, M. T.·Miyoshi, K.·Saito, K.
View original source ↗Export metadata ↓

Abstract

N{superscript 1}-methyladenosine at position 58 (m{superscript 1}A58), installed by the TRMT6/TRMT61A complex, is a highly conserved structural modification implicated in tRNA stability and human disease. In yeast, loss of m{superscript 1}A58 disrupts initiator methionine tRNA (tRNA) metabolism through defects in precursor processing and rapid tRNA decay, but whether these mechanisms operate in human cells has remained unclear. Here, using acute degron-mediated depletion of TRMT6 in human HCT116 cells combined with precursor-resolved tRNA sequencing, we show that m{superscript 1}A58 controls two distinct steps of tRNA metabolism. TRMT6 depletion selectively reduces mature tRNA while causing accumulation of precursor species retaining both 5'-leader and 3'-trailer sequences, consistent with impaired maturation. In parallel, the nuclear 5'[->]3' exonuclease XRN2 selectively degrades the residual mature hypomodified tRNA pool without affecting precursor accumulation. Loss of mature tRNA activates the integrated stress response and impairs proliferation, which is restored by XRN2 co-depletion. Together, our findings identify m{superscript 1}A58 as a coordinator of human initiator-tRNA maturation and surveillance, establishing a two-step pathway that maintains tRNA homeostasis in human cells.

Explore related subjects

biochemistryPreprint

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Otsubo, K., Nagura, M., Terauchi, M., Noguchi, H., Kanemaki, M. T., Miyoshi, K., Saito, K.. 2026-05-29. m1A58 governs two steps of human initiator-methionine tRNA metabolism: maturation and XRN2-mediated decay. https://doi.org/10.64898/2026.05.28.728624

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related preprints

bioRxiv2026

Conjunctive Targeting Links Drug Synergy to Emergent Proteome Structural States

Combinatorial therapies are widely used in the treatment of acute myeloid leukemia (AML) to address disease heterogeneity, adaptive resistance, and rewired signaling and metabolic states. Yet drug prioritization remains largely guided by clinical or phenotypic evidence, while the molecular mechanisms underlying effective drug combinations remain incompletely defined. To narrow this gap, we developed Combinatorial high-ratio Partial proteolysis with reference PRoteome Analysis (CoPPRA), a structural proteomics workflow based on limited proteolysis of cell lysates that profiles drug-associated changes in regional protein accessibility at peptide-level resolution. Here, we applied CoPPRA to ruxolitinib and ulixertinib, individually and in combination, in AML-related cell lysates. Our findings extend conjunctive targeting (CT), a recently proposed mechanism of combinatorial drug action in which combined exposure produces protein targeting patterns not observed with either drug alone. Previously identified through combination-associated changes in protein solubility/stability, CT is examined here at peptide-level resolution through regional differences in proteolytic accessibility. The ruxolitinib-ulixertinib combination produced broad peptide-level accessibility changes, including a subset meeting the predefined criteria for CT. CT candidates predominantly exhibited regional accessibility changes, with altered peptide regions occurring against comparatively small changes across the remaining quantified peptides from the same proteins. MAP2K1 and ATP6V1G1 showed pronounced differences between overlapping peptide sequences, highlighting localized variation in combination-associated accessibility, including an ATP6V1G1 peptide mapping to an annotated helical region. Combination-associated increases in peptide signals were also observed in PIK3R1, BRD4, and PTPN11, linking regional accessibility changes to signaling and transcriptional regulators relevant to AML. Functional enrichment and network analyses further implicated nucleotide and glucose metabolism, ficolin-1-rich granules, ribosome-associated processes, and phagocytic vesicles. These results extend conjunctive targeting from protein-level solubility/stability changes to regional differences in proteolytic accessibility, showing that combination-associated effects can be concentrated within specific peptide regions rather than distributed uniformly across proteins. More broadly, CoPPRA provides a peptide-resolved approach for investigating the molecular features of combinatorial drug action and prioritizing protein regions for subsequent mechanistic validation.

biochemistry↗
bioRxiv2026

Structural and biochemical characterisation of an iterative GCN5-related N-acetyltransferase required for fungal siderophore tailoring

Siderophore-mediated iron acquisition is essential for fungal survival, particularly under iron-limiting conditions. In Aspergillus fumigatus, SidG, a member of the GCN5-related N-acetyltransferase (GNAT) superfamily, catalyses the final step in the biosynthesis of the extracellular siderophore triacetylfusarinine C (TAFC) through sequential acetylation of the precursor fusarinine C (FsC). However, the timing, catalytic mechanism, and functional significance of this modification are not fully understood. Here, we reconstituted SidG activity in vitro and combined native mass spectrometry, X-ray crystallography, molecular dynamics simulations, and site-directed mutagenesis to investigate its catalytic properties. Our analyses demonstrate that SidG selectively binds acetyl-CoA from the cellular milieu and iteratively acetylates the FsC scaffold prior to iron chelation. Structural, biochemical, and molecular dynamics analyses support a direct transfer mechanism, identify key catalytic residues, and demonstrate the strict selectivity of SidG for short-chain acyl-CoA donors. Together, these findings establish the molecular basis for SidG-dependent siderophore tailoring and expand our understanding of GNAT-catalysed transformations in fungal natural product biosynthesis.

biochemistry↗
bioRxiv2026

Reconstitution of +1 nucleosome transcription reveals coordinated functions of SAGA, Mediator, and TFIIH

The +1 nucleosome has emerged as a key regulator of eukaryotic transcription, but how it controls transcription initiation remains poorly understood. Here we reconstitute transcription through the +1 nucleosome using eleven purified yeast factors: RNA polymerase II (Pol II), the six general transcription factors (GTFs), TFIIS, the activator Pho4, and the SAGA and Mediator complexes. The system recapitulates key features of regulation observed in vivo. SAGA, acting with Pho4, directs pre-initiation complex (PIC) assembly to the correct position through its TBP-loading activity. Mediator stimulates transcription when the +1 nucleosome imposes a barrier to PIC formation, consistent with stabilization of productive TFIIH-DNA engagement. Contrary to the prevailing model, SAGA remains bound to the PIC after TBP loading and acetylates the +1 nucleosome within the assembled complex. The isolated PIC-Mediator-SAGA-nucleosome complex is transcriptionally active, and the repressive effect of the nucleosome is relieved by the DNA translocase activity of Ssl2, the TFIIH subunit that opens promoter DNA. TFIIH thus couples promoter melting to remodeling of the +1 nucleosome.

biochemistry↗