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Regulation of Expression at Multiple Steps01:23

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The gene expression in cells is regulated at different stages: (i) transcription, (ii) RNA processing, (iii) RNA localization, and (iv) translation. Transcriptional regulation is mediated by regulatory proteins such as transcription factors, activators, or repressors—these control gene expression by initiating or inhibiting the transcription of genes. Once a precursor or pre-mRNA is produced, it undergoes post-transcriptional modification, including 5' capping, splicing, and the...
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Multimodal profiling reveals cell type-specific pseudouridine modification and density-dependent translational

Caroline A McCormick1, Michele Meseonznik1, Yuchen Qiu1

  • 1Dept. of Bioengineering, Northeastern University, Boston, MA, 02115, United States.

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Pseudouridine (psi) single-site modification enhances protein production, while high-density pseudouridylation impairs translation. This study reveals pseudouridine density and enzyme specificity as key determinants of proteome output.

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Area of Science:

  • Molecular Biology
  • Epigenetics
  • Proteomics

Background:

  • Pseudouridine (psi) is a prevalent mRNA modification with an unclear role in translation.
  • Existing pseudouridine maps are inconsistent, hindering cross-cell type comparisons and translation studies.

Purpose of the Study:

  • To map pseudouridine modifications at single-nucleotide resolution across human cell lines.
  • To investigate the impact of pseudouridine on protein production and translation efficiency.
  • To elucidate the mechanistic role of pseudouridine density and enzyme specificity in regulating proteome output.

Main Methods:

  • Direct RNA nanopore sequencing for high-resolution pseudouridine mapping.
  • Mod-p ID analytical framework for pseudouridine quantification and co-modification detection.
  • Integration with proteomic and ribosome profiling data, including TRUB1 knockout experiments and in vitro translation assays.

Main Results:

  • Conserved pseudouridine sites, particularly those installed by TRUB1, correlate with increased protein production.
  • TRUB1 knockout confirms pseudouridylation's causal role in enhancing protein output in a motif-specific manner.
  • Clustered pseudouridine sites reduce protein abundance, despite increased translation efficiency, demonstrating a density-dependent inhibitory effect.

Conclusions:

  • Single-site pseudouridylation enhances protein production, whereas hypermodification impairs translational throughput.
  • Pseudouridine density and specific enzyme action (e.g., TRUB1) are critical regulators of proteome output.
  • This study provides a mechanistic framework for understanding pseudouridine's dual role in translation across human cell types.