Transcriptome-wide quantitative profiling of PUS7-dependent pseudouridylation by nanopore direct long read RNA sequencing

Bansal, M.; Kundu, A.; Gibson, A.; Gupta, A.; Ding, J.; RudraRaju, S. V.; Sudarshan, S.; Ding, H.-F.

doi:10.1101/2024.01.31.578250

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bioRxiv · 10.1101/2024.01.31.578250

Transcriptome-wide quantitative profiling of PUS7-dependent pseudouridylation by nanopore direct long read RNA sequencing

Bansal, M.·Kundu, A.·Gibson, A.·Gupta, A.·Ding, J.·RudraRaju, S. V.·Sudarshan, S.·Ding, H.-F.

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Abstract

Understanding the interactions between different RNA modifications is essential for unraveling their biological functions. Here, we report NanoPsiPy, a computational pipeline that employs nanopore direct RNA sequencing to identify pseudouridine ({Psi}) sites and quantify their levels at single-nucleotide resolution. We validated NanoPsiPy by transcriptome-wide profiling of PUS7-dependent {Psi} sites in poly-A RNA and rRNA. NanoPsiPy leverages {Psi}-induced U-to-C basecalling errors in nanopore sequencing data, allowing detection of both low and high stoichiometric {Psi} sites. We identified 8,624 PUS7-dependent {Psi} sites in 1,246 mRNAs encoding proteins associated with ribosome biogenesis, translation, and energy metabolism. Importantly, integrative analysis revealed that PUS7 knockdown increases global mRNA N6-methyladenosine (m6A) and 5-methylcytosine (m5C) levels, suggesting an antagonistic relationship between {Psi} and these modifications. Our study underscores the potential of nanopore direct RNA sequencing in revealing the co-regulation of RNA modifications and the capacity of NanoPsiPy in analyzing pseudouridylation and its impact on other RNA modifications.

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Bansal, M., Kundu, A., Gibson, A., Gupta, A., Ding, J., RudraRaju, S. V., Sudarshan, S., Ding, H.-F.. 2024-02-01. Transcriptome-wide quantitative profiling of PUS7-dependent pseudouridylation by nanopore direct long read RNA sequencing. https://doi.org/10.1101/2024.01.31.578250

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