NAD-RNA Decapping in Escherichia coli: Discovering the Mechanism and Function of NudC
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Philipps-Universität Marburg
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Abstract
In Escherichia coli, NudC plays a crucial role in the decapping of 5’-NAD-RNA. Initially identified as a Nudix hydrolase capable of hydrolyzing NAD+ into adenosine monophosphate (AMP) and nicotinamide mononucleotide (NMN), recent studies have shown that NudC has a stronger substrate preference for 5’-NAD-RNA over NAD+. This preference results in the decapping of 5’-NAD-RNA, producing 5’-P-RNA and NMN. The association between NudC and RNA was first observed during protein purification in 1995, but this interaction remained unexplored until recent next-generation sequencing (NGS) data revealed that the RNA binding by NudC is independent of RNA size and sequence. However, the mechanism in which NudC binds to RNA substrates and the biological implications that this interaction might regulate, remain unexplored. Therfore, this work aims to extend the knowledge gap of NAD-RNA decapping mediated by NudC and further implications of it in E. coli.
Hereby we present an extensive structural, biochemical, and phenotypical analysis demonstrating the mechanism in which NudC decapping activity, localization, and protein interactions are mediated by RNA binding.
Therefore, in section 3.1 we identified by an in silico approach the critical residues of NudC involved in RNA interaction. By in vitro data demonstrated that residues R120 and R122 mediate NudC-RNA binding, which facilitates the decapping of NAD-RNA. By in vivo assays, we demonstrated that the absence of these residues increase up to 10 times the half-life of NAD-RNAs transcripts. Motivated by these results, in section 3.2 we demonstrated that Nud-RNA binding is crucial to promote NudC localization and diffusion towards the inner membrane, and intracellular RNA drives such behavior.
Additionally, in section 3.3 we showed for the first time by in vivo and in silico evidence, the association between NudC and the RNA polymerase complex, proposing that decapping and transcription, are coupled processes in E. coli. We also identified the interaction between NudC and PNPase proposing a novel 3’ dependent NAD-RNA degradation by PNPase facilitated by NudC-RNA binding. Ultimately, in section 3.4 we identified protein dysregulation upon NudC-RNA binding disruption, which leads to impaired Biofilm formation in E. coli, possibly linked to untranslated NAD-mRNAs.
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Created: 2025Issued: 2026-01-12Updated: 2025-07-17
Faculty
Fachbereich Chemie
Publisher
Philipps-Universität Marburg
Language
eng
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DoctoralThesis
Keywords
Bacterial EpitranscriptomicsRNA biology
DFG-subjects
MicrobiologyMolecular Biology.
DDC-Numbers
570
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Rodriguez Mendez, David: NAD-RNA Decapping in Escherichia coli: Discovering the Mechanism and Function of NudC. : Philipps-Universität Marburg 2026-01-12. DOI: https://doi.org/10.17192/z2025.0506.