The proximity proteome of pre-40S pre-ribosomal particle components PNO1 and NOB1 using turboID proximity labeling technology

Abstract

Background

The ribosome assembly factors PNO1 and NOB1 play crucial roles in the maturation of the 40S ribosomal small subunit. TurboID is an efficient biotin ligase that can biotinylate proteins in proximity to the target protein and is widely used to study complex biological processes within cells. In this study, we employed this technology to investigate the complex proximity network of PNO1 and NOB1 within the cell, further revealing their key roles in ribosome biogenesis.

Results

Firstly, through immunofluorescence experiments, we observed that PNO1 and NOB1 have different localizations within the cell. Subsequently, by analyzing the proximal proteins labeled by PNO1-TurboID and NOB1-TurboID, we identified 871 proximal proteins for PNO1 and 1044 for NOB1, with 663 overlapping proteins. Functional analysis revealed that these proximal proteins are predominantly enriched in biological processes related to ribosome assembly, rRNA processing, and translation, all of which are closely linked to ribosome biogenesis. Notably, we validated the mass spectrometry-identified proteins through co-IP experiments and found that PNO1 and NOB1 interact with the translation-related proteins EIF4B and EIF4G2.

Conclusion

Our study constructed the protein network environment of ribosome assembly factors PNO1 and NOB1 within the cell and found that their neighboring proteins are primarily involved in key biological processes such as ribosome processing, mRNA translation, and the cell cycle, all of which are critical for ribosome biogenesis. These findings provide a valuable foundation for further understanding the roles of PNO1 and NOB1 in ribosome biogenesis and how they regulate this process.