Opportunities and Challenges to Profile mRNA Modifications in Escherichia coli

2022-07-13

Dimitar Plamenov Petrov, Steffen Kaiser, Stefanie Kaiser, and Kirsten Jung

ChemBioChem 2022, 23, e202200270

https://doi.org/10.1002/cbic.202200270

mRNA methylation is an important regulator of many physiological processes in eukaryotes but has not been studied in depth in prokaryotes. Working with bacterial mRNA is challenging because it lacks a poly(A)-tail. However, methods for detecting RNA modifications, both sequencing and mass spectrometry, rely on efficient preparation of mRNA. Here, we compared size-dependent separation by electrophoresis and rRNA depletion for enrichment of Escherichia coli mRNA. The purification success  as monitored by qRT-PCR and RNA sequencing. Neither method allowed complete removal of rRNA. Nevertheless, we were able to quantitatively analyze several modified nucleosides in the different RNA types. We found evidence for stress dependent RNA modification reprofiling in rRNA, but also several modified nucleosides in the mRNA enriched fractions showed significant changes.