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Article Dans Une Revue RNA Année : 2023

Nanopore-based RNA sequencing deciphers the formation, processing, and modification steps of rRNA intermediates in archaea

Felix Grünberger
Michael Jüttner
  • Fonction : Auteur
Robert Knüppel
  • Fonction : Auteur
Dina Grohmann

Résumé

Ribosomal RNA (rRNA) maturation in archaea is a complex multistep process that requires well-defined endo- and exoribonuclease activities to generate fully mature linear rRNAs. However, technical challenges prevented detailed mapping of rRNA processing steps and a systematic analysis of rRNA maturation pathways across the tree of life. In this study, we used long-read (PCR)-cDNA and direct RNA nanopore-based sequencing to study rRNA maturation in three archaeal model organisms, namely the Euryarchaea Haloferax volcanii and Pyrococcus furiosus and the Crenarchaeon Sulfolobus acidocaldarius . Compared to standard short-read protocols, nanopore sequencing facilitates simultaneous readout of 5′- and 3′-positions, which is required for the classification of rRNA processing intermediates. More specifically, we (i) accurately detect and describe rRNA maturation stages by analysis of terminal read positions of cDNA reads and thereupon (ii) explore the stage-dependent installation of the KsgA-mediated dimethylations in H. volcanii using base-calling and signal characteristics of direct RNA reads. Due to the single-molecule sequencing capacity of nanopore sequencing, we could detect hitherto unknown intermediates with high confidence, revealing details about the maturation of archaea-specific circular rRNA intermediates. Taken together, our study delineates common principles and unique features of rRNA processing in euryarchaeal and crenarchaeal representatives, thereby significantly expanding our understanding of rRNA maturation pathways in archaea.
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Origine : Publication financée par une institution
Licence : CC BY NC - Paternité - Pas d'utilisation commerciale

Dates et versions

hal-04207618 , version 1 (17-10-2023)

Identifiants

Citer

Felix Grünberger, Michael Jüttner, Robert Knüppel, Sébastien Ferreira-Cerca, Dina Grohmann. Nanopore-based RNA sequencing deciphers the formation, processing, and modification steps of rRNA intermediates in archaea. RNA, 2023, 29 (8), pp.1255-1273. ⟨10.1261/rna.079636.123⟩. ⟨hal-04207618⟩
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